Use of hexokinase 2/mitochondria-detaching compounds for activating immune responses

ABSTRACT

The present invention provides methods of use of hexokinase 2 (HK2)/mitochondria-detaching compounds, including jasmonate derivatives and piperazine derivatives and pharmaceutical compositions including such compounds for inducing immune responses in a subject, including potentiating the immune response to hyperproliferative disorders such as cancer and potentiating the immune response to infectious diseases.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a Divisional application of U.S. patent applicationSer. No. 15/576,824 filed Nov. 26, 2017 which is a National PhaseApplication of PCT International Application No. PCT/IL2017/051213 filedNov. 7, 2017, which claims priority to U.S. Provisional Application No.62/418,323, filed Nov. 7, 2016, and to U.S. Provisional Application No.62/573,179, filed Oct. 17, 2017, which are hereby incorporated byreference in their entirety.

FIELD OF THE INVENTION

The present invention provides methods of use of hexokinase 2(HK2)/mitochondria-detaching compounds, including jasmonate derivativesand piperazine derivatives and pharmaceutical compositions includingsuch compounds for potentiating immune responses in a subject, includingpotentiating immune responses in subjects with hyperproliferativedisorders such as cancer and potentiating the immune response toinfectious diseases.

BACKGROUND OF THE INVENTION

There is evidence to suggest the immune system may inhibit thedevelopment of cancer, via the innate immune system and/or the adaptiveimmune system, possibly via cancer-associated recognition events. Thehost may have a dedicated mechanism to perceive and eliminatetransformed cells. In addition, adaptive immune recognition oftumor-associated and specific antigens also may be an important means bywhich the immune system controls the development of cancer. Furthermore,memory immune cells have an important role in anti-cancerimmune-surveillance. Compounds that can induce an immune response insubjects with a hyperproliferative disorder are lacking.

SUMMARY OF THE INVENTION

In one embodiment, the present invention provides a compound thatpromotes the detachment of Hexokinase 2 (HK2) from the mitochondria foruse in inducing an immune response in a subject.

In another embodiment, the present invention provides a compound thatpromotes the detachment of Hexokinase 2 (HK2) from the mitochondria foruse in potentiating the immune response to a hyperproliferative disorderin a subject.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. Efficacy of topical Formula C treatment on tumor volume in themouse B16-F10 melanoma model. * p≤0.05, *** p≤0.001.

FIG. 2. Topical Formula C administration increases inflammation in themouse B16-F10 melanoma model in the ear. * p≤0.05.

FIG. 3A. Box and Whiskers plot of semi-quantitative analysis of positiveimmunohistochemistry staining of skin sections stained with toluidineblue for mast cells following 50-day of topical Formula C treatment ofUVB-exposed mice. Quantification included the number of positive cellsper x40 high power field, 7 different fields per slide.

FIG. 3B. Box and Whiskers plot of semi-quantitative analysis of positiveimmunohistochemistry staining of skin sections stained with CD3⁺ for Tcells following 50-day of topical Formula C treatment of UVB-exposedmice. Quantification included the number of positive cells per x40 highpower field, 7 different fields per slide.

FIG. 3C. Box and Whiskers plot of semi-quantitative analysis of positiveimmunohistochemistry staining of skin sections stained with Langerin forskin dendritic cells following 50-day of topical Formula C treatment ofUVB-exposed mice. Quantification included the number of positive cellsper x40 high power field, 7 different fields per slide.

FIG. 4. Formula C stimulates IL-1β secretion in activated humanmacrophages in vitro. THP-1 human monocytes were differentiated tomature macrophages and stimulated with LPS (Lipopolysaccharide) followedby Formula C treatment. Secreted IL-1 β was analyzed in the culturemedium by ELISA.

FIG. 5A. Formula C dose-dependently stimulates IL-1β secretion inprimary bone marrow-derived macrophages from Balb/C mouse. SecretedIL-1β was analyzed in the culture medium by ELISA.

FIG. 5B. The effect of Formula C treatment on the viability of primarybone marrow-derived macrophages from Balb/C mouse.

FIG. 5C. Formula C dose-dependently stimulates IL-1β secretion inprimary bone marrow-derived macrophages from C57BL mouse. Secreted IL-1β was analyzed in the culture medium by ELISA. UT, untreated. HK(hexokinase-derived peptide) serves as a positive control.

FIG. 5D. The effect of Formula C treatment on the viability of primarybone marrow-derived macrophages from C57BL mouse. UT, untreated. HK(hexokinase peptide) serves as a positive control.

FIG. 6A. Formula C administration decreased tumor volume in Balb/C micein syngeneic cancer model (CT26 mouse colorectal cancer).

FIG. 6B. Formula C administration shifts macrophages in vivo frompro-tumor (M2) macrophages to antitumor (M1) macrophages. CT26 tumorsfrom Formula C or vehicle-treated Balb/C mice with CT26 mouse colorectalcancer was analyzed for MHCII marker by FACS. Dark color: Vehicletreated CT26 tumors from Balb/C mice (day 20). Light color: Formula C(150 mg/kg) treated CT26 tumors from Balb/C mice (day 20).

FIG. 6C. Formula C administration shifts macrophages in vivo frompro-tumor (M2) macrophages to antitumor (M1) macrophages. CT26 tumorsfrom Formula C or vehicle-treated Balb/C mice with CT26 mouse colorectalcancer was analyzed for CD206. Dark color: Vehicle treated CT26 tumorsfrom Balb/C mice (day 20). Light color: Formula C (150 mg/kg) treatedCT26 tumors from Balb/C mice (day 20).

FIGS. 7A-F. Formula C administration shifts effector CD8+ T cells tocentral memory cells. Primary Balb/C mouse splenocytes were grown inculture and activated by anti-CD3e and anti CD28 antibodies to shiftthem towards CD8+/CD4+ phenotype. Cells were then treated with Formula C(FIG. 7E), 2-DG (2-deoxyglucose, positive control) (FIG. 7C), or vehicle(FIG. 7B, FIG. 7D) for 24 hours followed by FACS analysis for CD44 andCD62L. FIG. 7A shows FACs analysis for CD44 and CD62L in non-activatedsplenoctyes. Formula C administration did not reduce the percentage oflive cells in the sample (FIG. 7F).

DETAILED DESCRIPTION OF THE PRESENT INVENTION

In the following detailed description, numerous specific details are setforth in order to provide a thorough understanding of the invention.However, it will be understood by those skilled in the art that thepresent invention may be practiced without these specific details. Inother instances, well-known methods, procedures, and components have notbeen described in detail so as not to obscure the present invention.

In one embodiment, the present invention provides a method of inducingan immune response in a subject comprising the step of contacting one ormore cells of said subject with a therapeutically effective amount of acompound that promotes the detachment of Hexokinase 2 (HK2) from themitochondria. In one embodiment, the compound is a jasmonate derivative.In another embodiment, the compound is a piperazine derivative.

In another embodiment, the present invention provides a method ofactivating an immune response in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of a compound that promotes the detachment ofHexokinase 2 (HK2) from the mitochondria. In one embodiment, thecompound is a jasmonate derivative. In another embodiment, the compoundis a piperazine derivative.

In another embodiment, the present invention provides a method ofpotentiating an immune response in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of a compound that promotes the detachment ofHexokinase 2 (HK2) from the mitochondria. In one embodiment, thecompound is a jasmonate derivative. In another embodiment, the compoundis a piperazine derivative.

In one embodiment, the compounds as described herein may be used toactivate anti-cancer immune responses. In another embodiment, thecompounds as described herein may be used to potentiate anti-cancerimmune responses.

Surprisingly and unexpectedly, the inventors have found thatadministration of the methyl jasmonate derivative Formula C leads toincreased inflammatory response (FIG. 2) and the activation of an immuneresponse in the skin of UVB-exposed mice model of actinic keratosis andsquamous cell carcinoma (FIGS. 3A-C). Formula C administration alsoresulted in increased IL-1β secretion from macrophages (FIGS. 4 and5A-D), and a shift of macrophages from the pro-tumor M2 phenotype to ananti-tumor M1 phenotype (FIGS. 6A-C). Finally, Formula C administrationshifts T cells from effector T cells to memory T cells (FIGS. 7A-F).

Immune Responses

As used herein, the term “immune response” refers to any detectableresponse by the immune system of a host vertebrate animal, including,but not limited to, innate immune responses (e.g., activation of Tollreceptor signaling cascade and stimulation of macrophages),cell-mediated immune responses (e.g., responses mediated by T cells,such as antigen-specific T cells, and non-specific cells of the immunesystem), and humoral immune responses (e.g., responses mediated by Bcells, such as generation and secretion of antibodies into the plasma,lymph, and/or tissue fluids). Examples of immune responses include analteration (e.g., increase) in Toll-like receptor activation, lymphokine(e.g., cytokine (e.g., Th1, Th2 or Th17 type cytokines) or chemokine)expression or secretion, macrophage activation, stimulation ofanti-tumor M1 macrophage phenotype, secretion of cytokines frommacrophages, dendritic cell activation, T cell (e.g., CD4+ or CD8+Tcell) activation, induction of memory T cells, NK cell activation, Bcell activation (e.g., antibody generation and/or secretion), mast cellactivation, binding of an immunogen (e.g., antigen (e.g., immunogenicpolypolypeptide)) to an MHC molecule, induction of a cytotoxic Tlymphocyte (“CTL”) response, induction of a B cell response (e.g.,antibody production), and expansion (e.g., growth of a population ofcells) of cells of the immune system (e.g., T cells and B cells), andincreased processing and presentation of antigen by antigen presentingcells. The term “immune response” also encompasses any detectableresponse to a particular substance (such as an antigen or immunogen) byone or more components of the immune system of a vertebrate animal invitro.

In one embodiment, “inducing an immune response” may mean that an immuneresponse was induced in a situation where there was no prior immuneresponse. In another embodiment, an immune response is enhanced in asituation where there was some level of immune response before immuneresponse induction. Thus, “inducing an immune response” includes“enhancing an immune response”. In another embodiment, an inactivecomponent of the immune system (e.g., macrophages or T cells) isactivated in addition to other already activated components of theimmune system. In another embodiment, the activity of an activecomponent of the immune system (e.g., macrophages or T cells) isenhanced in addition to other already activated components of the immunesystem. In another embodiment, the activity of a component of the immunesystem (e.g., macrophages or T cells) is enhanced while the activity ofanother components of the immune system is decreased. Preferably, afterinducing an immune response in a subject, said subject is protected fromdeveloping a disease or disorder such as cancer or the disease ordisorder is ameliorated by inducing an immune response.

Thus, in one embodiment, the present invention provides a method ofinducing a humoral immune response in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of a HK2/mitochondria-detaching compound. In anotherembodiment, the present invention provides a method of inducing aninnate immune response in a subject comprising the step of contactingone or more cells of said subject with a therapeutically effectiveamount of a HK2/mitochondria-detaching compound. In another embodiment,the present invention provides a method of inducing a cell-mediatedimmune response in a subject comprising the step of contacting one ormore cells of said subject with a therapeutically effective amount of aHK2/mitochondria-detaching compound. In one embodiment, theHK2/mitochondria-detaching compound is a jasmonic acid derivative. Inanother embodiment, the HK2/mitochondria-detaching compound is apiperazine derivative.

Thus, in one embodiment, the present invention provides a method ofactivating a humoral immune response in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of a HK2/mitochondria-detaching compound. In anotherembodiment, the present invention provides a method of activating aninnate immune response in a subject comprising the step of contactingone or more cells of said subject with a therapeutically effectiveamount of a HK2/mitochondria-detaching compound. In another embodiment,the present invention provides a method of activating a cell-mediatedimmune response in a subject comprising the step of contacting one ormore cells of said subject with a therapeutically effective amount of aHK2/mitochondria-detaching compound. In one embodiment, theHK2/mitochondria-detaching compound comprises a jasmonic acidderivative. In one embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula (VII) as describedherein. In another embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula C, as described herein.In another embodiment, the HK2/mitochondria-detaching compound comprisesa piperazine derivative.

Thus, in one embodiment, potentiating a humoral immune response in asubject comprising the step of contacting one or more cells of saidsubject with a therapeutically effective amount of anHK2/mitochondria-detaching compound. In another embodiment, the presentinvention provides a method of potentiating an innate immune response ina subject comprising the step of contacting one or more cells of saidsubject with a therapeutically effective amount of anHK2/mitochondria-detaching compound. In another embodiment, the presentinvention provides a method of potentiating a cell-mediated immuneresponse in a subject comprising the step of contacting one or morecells of said subject with a therapeutically effective amount of anHK2/mitochondria-detaching compound. In one embodiment, theHK2/mitochondria-detaching compound comprises a jasmonic acidderivative. In one embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula (VII) as describedherein. In another embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula C, as described herein.In another embodiment, the HK2/mitochondria-detaching compound comprisesa piperazine derivative.

In one embodiment, the present invention provides a method of activatingT cells in a subject comprising the step of contacting one or more cellsof said subject with a therapeutically effective amount of aHK2/mitochondria-detaching compound. In another embodiment, the T cellactivation is enhanced T cell proliferation, enhanced cytokineproduction and/or enhanced T cell expression. In one embodiment, theHK2/mitochondria-detaching compound comprises a jasmonic acidderivative. In one embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula (VII) as describedherein. In another embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula C, as described herein.In another embodiment, the HK2/mitochondria-detaching compound comprisesa piperazine derivative.

In one embodiment, the term “T cell” refers to T lymphocytes as definedin the art and is intended to include thymocytes, immature Tlymphocytes, mature T lymphocytes, resting T lymphocytes, or activated Tlymphocytes. The T cells can be CD4+ T cells, CD8+ T cells, CD4+ CD8+ Tcells, or CD4−CD8− cells. The T cells can also be T helper cells, suchas T helper 1 (TH1), or T helper 2 (TH2) cells, or TH17 cells, as wellas cytotoxic T cells, regulatory T cells, natural killer T cells, naïveT cells, memory T cells, or gamma delta T cells.

In another embodiment, the present invention provides a method ofreprogramming T cells to memory T cells. In another embodiment, thepresent invention provides a method of converting T cells to memory Tcells. In another embodiment, the present invention provides a method ofshifting T cells to memory T cells. In another embodiment, the presentinvention provides a method of increasing the number or percentage ofmemory T cells. In another embodiment, the present invention provides amethod of increasing or expediting the effector to memory T celltransition.

In another embodiment, the present invention provides a method ofincreasing CD62L levels, CD44 levels, or a combination thereof in Tcells, comprising the step of contacting one or more cells of saidsubject with a therapeutically effective amount of anHK2/mitochondria-detaching compound. In one embodiment, theHK2/mitochondria-detaching compound comprises a jasmonic acidderivative. In one embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula (VII) as describedherein. In another embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula C, as described herein.In another embodiment, the HK2/mitochondria-detaching compound comprisesa piperazine derivative.

In one embodiment, memory T cells are central memory T cells (T_(CM)cells). In another embodiment, memory T cells are effector memory Tcells (TEM cells). In another embodiment, memory T cells are stem memoryT cells (T_(SCM) cells). In one embodiment, memory cells may be eitherCD4+ or CD8+.

In one embodiment, the methods as described herein are performed invivo. In another embodiment, the methods are performed in vitro. In oneembodiment, the methods are performed on primary cells. In oneembodiment, the T cells can be from previously stored blood samples,from a healthy individual, or alternatively from an individual affectedwith a condition. In one embodiment, the condition is ahyperproliferative disease. In another embodiment, the condition is aninfectious disease, such as a condition resulting from a viralinfection, a bacterial infection or an infection by any othermicroorganism. In one embodiment, the T cells can be of human origin,murine origin or any other mammalian species.

In another embodiment, the present invention provides a method ofconverting macrophages from pro-tumor M2 to anti-tumor M1 phenotypecomprising the step of contacting one or more cells of said subject witha therapeutically effective amount of a hexokinase 2(HK2)/mitochondria-detaching compound. In another embodiment, thepresent invention provides a method of increasing MHCII levels,decreasing CD206 levels, or a combination thereof in a macrophage,comprising the step of contacting one or more cells of said subject witha therapeutically effective amount of an HK2/mitochondria-detachingcompound. In one embodiment, the HK2/mitochondria-detaching compoundcomprises a jasmonic acid derivative. In one embodiment, the jasmonicacid derivative comprises a compound represented by the structure ofFormula (VII) as described herein. In another embodiment, the jasmonicacid derivative comprises a compound represented by the structure ofFormula C, as described herein. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative.

In another embodiment, the present invention provides a method ofstimulating secretion of cytokines from macrophages, comprising the stepof contacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In one embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula (VII) asdescribed herein. In another embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula C, asdescribed herein. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative. In one embodiment, thecytokine comprises IL-1β, IL-18, or a combination thereof.

In another embodiment, the present invention provides a method ofactivating NK cells in a subject comprising the step of contacting oneor more cells of said subject with a therapeutically effective amount ofan HK2/mitochondria-detaching compound. In one embodiment, the NK cellactivation is enhanced NK cell proliferation, enhanced cytokineproduction and/or enhanced NK cell expression. In another embodiment,the present invention provides a method of activating dendritic cells ina subject comprising the step of contacting one or more cells of saidsubject with a therapeutically effective amount of anHK2/mitochondria-detaching compound. In another embodiment, thedendritic cells activation is enhanced dendritic cells proliferation,enhanced cytokine production and/or enhanced dendritic cells expression.In another embodiment, the present invention provides a method ofactivating mast cells in a subject comprising the step of contacting oneor more cells of said subject with a therapeutically effective amount ofan HK2/mitochondria-detaching compound. In another embodiment, the mastcells activation is enhanced mast cells proliferation, enhanced cytokineproduction and/or enhanced mast cells expression. In another embodiment,the present invention provides a method of lowering the levels ofmyeloid-derived suppressor cells (MDSCs) comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In one embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula (VII) asdescribed herein. In another embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula C, asdescribed herein. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative.

In one embodiment, the present invention provides a method of inducinginflammation in a subject comprising the step of contacting one or morecells of said subject with a therapeutically effective amount of anHK2/mitochondria-detaching compound. In one embodiment, the inflammationis acute inflammation. In another embodiment, the inflammation ischronic inflammation. In one embodiment, the HK2/mitochondria-detachingcompound comprises a jasmonic acid derivative. In one embodiment, thejasmonic acid derivative comprises a compound represented by thestructure of Formula (VII) as described herein. In another embodiment,the jasmonic acid derivative comprises a compound represented by thestructure of Formula C, as described herein. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative.

In another embodiment, the present invention provides a method ofactivating an inflammatory response in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In one embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula (VII) asdescribed herein. In another embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula C, asdescribed herein. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative.

In another embodiment, the present invention provides a method ofactivating an inflammasome in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In one embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula (VII) asdescribed herein. In another embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula C, asdescribed herein. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative.

In another embodiment, the present invention provides a method ofinflammasome activation in a subject comprising the steps as describedherein. In another embodiment, the present invention provides a methodof inflammasome regulation in a subject comprising the steps asdescribed herein. In another embodiment, the present invention providesa method of activating an inflammasome-mediated immune response in asubject comprising the steps as described herein.

Hexokinase 2 (HK2)/Mitochondria-Detaching Compounds

In one embodiment, compounds for use in the methods of the presentinvention comprise compounds that promote the detachment of HK2 from themitochondria. In another embodiment, compounds for use in the methods ofthe present invention comprise compounds that promote the detachment ofHK2 from VDAC (voltage-dependent anion channel). In another embodiment,compounds for use in the methods of the present invention comprisecompounds that disrupt the physical contact between HK2 and themitochondria in general or specifically between HK2 and VDAC.

In one embodiment, compounds that disrupt HK2/mitochondria attachmentare known in the art. In another embodiment, compounds may be evaluatedfor such function by a skilled artisan. In one embodiment, disruption ofHK2/mitochondria attachment may be assayed in a cell-free environment,using a technology such as microscale thermophoresis (MST), or othersevaluating protein-protein interaction modulators (as described in U.S.Provisional Patent Application No. 62/577,256, which is incorporated byreference herein in its entirety). In another embodiment, disruption ofHK2/mitochondria attachment may be assayed in a cell-based assay usingvarious methods including, inter alia, Western blot, wherein themitochondrial fraction is examined. Such methods are well known in theart.

In one embodiment, the compound comprises methyl jasmonate. In anotherembodiment, the compound comprises a methyl jasmonate derivative.

Jasmonate Derivatives

In one embodiment, the jasmonate derivative comprises a compoundrepresented by the structure of Formula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof.

In a further embodiment, the compound is represented by the structure offormula C:

In one embodiment, the structure of formula C decouples HK2 fromVoltage-dependent anion channel (VDAC) in cancer cells. In anotherembodiment, administration of an HK2/mitochondria-detaching compound toa cell leads to activation of an inflammasome in the cell. In anotherembodiment, administration of an HK2/mitochondria-detaching compound toa cell leads to an increase in one or more cytokines, which, in oneembodiment, comprise IL-1β, and, in another embodiment, IL-18. Inanother embodiment, administration of an HK2/mitochondria-detachingcompound to a cell leads to activation of an immune response in thecell. In one embodiment, the cell has high HK-2 expression. In oneembodiment, the cell is a cancer cell. In one embodiment, theHK2/mitochondria-detaching compound comprises a jasmonic acidderivative. In one embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula (VII) as describedherein. In another embodiment, the jasmonic acid derivative comprises acompound represented by the structure of Formula C, as described herein.In another embodiment, the HK2/mitochondria-detaching compound comprisesa piperazine derivative.

In another embodiment, A is COR¹; R¹ is quinolinyloxy; R² is oxo; R³,R⁴, R⁵, R⁶ and R⁷ are each hydrogen, wherein the bond between C₉ and C₁₀can be a single or double bond; R¹⁵ and R¹⁶ are each hydrogen; and n isselected from 0, 1 and 2; including salts, hydrates, solvates,polymorphs, optical isomers, geometrical isomers, enantiomers,diastereomers, and mixtures thereof.

In another embodiment, the compound is represented by the structure offormula C:

In one embodiment, ester derivatives of Jasmonic Acid may be used in themethods of the present invention. In another embodiment, other jasmonatederivatives may be used in the methods of the present invention. In oneembodiment, halogenated jasmonate derivatives and related pharmaceuticalcompositions, as described in International Patent Application WO2005/054172, which is incorporated by reference herein in its entirety,may be used in the methods of the present invention.

In another embodiment, jasmonate derivatives such as those described inInternational Patent Applications WO 2007/066336, WO 2010/143180, and WO2007/066337, which are incorporated by reference herein in theirentirety may be used in the methods of the present invention. In anotherembodiment, dermal compositions comprising jasmonate derivatives may beused in the methods of the present invention. Methods for preparing thejasmonate derivatives for use in the present invention are described,for example, in WO 2007/066336 and WO 2010/143180.

Piperazine Derivatives

In another embodiment, the compound for use in the present inventioncomprises piperazine derivatives, which in one embodiment, comprise thecompounds described in PCT Application No. PCT/IL2017/050909, which isincorporated herein by reference in its entirety. According to thisaspect and in one embodiment, the present invention provides the use ofa compound represented by the structure of Formula (II):

wherein

-   -   U and Z are each independently N or CH;    -   X is O, NH, S, or a bond;    -   Y is CH₂, C═O, or C═S;    -   R¹ is aryl, heteroaryl, or C(═O)—OR^(a), wherein aryl and        heteroaryl are each optionally substituted with one or more        alkyl, arylalkyl, halogen, NO₂, CN, OR⁴, NR^(5a)R^(5b), or a        combination thereof;    -   R² and R³ are each independently at each occurrence selected        from the group consisting of: halogen, NO₂, CN, C₁-C₄ alkyl,        OR⁴, and NR^(5a)R^(5b);    -   R⁴, R^(a), R^(5a), and R^(5b) are each independently hydrogen or        C₁-C₄ alkyl;    -   n is 0 or 1;    -   m is 0, 1, or 2; and    -   p and q are each independently selected from 0, 1, 2, 3, and 4;        or a pharmaceutically acceptable salt thereof, with the proviso        that when Z is CH, X is NH, S, or a bond.

In another embodiment, the following compound is excluded: a compound ofFormula (II) wherein X is NH, Z is CH, Y is C═O, n is 0, m is 0, R¹ is aphenyl substituted by one or more alkoxy, and p and q are each 0.

In some embodiments, in the compound of Formula (II), m and n are each0.

In some embodiments, in the compound of Formula (II), R¹ is aryl orheteroaryl. In some embodiments, R¹ is phenyl, quinolinyl, orisoquinolinyl. In other embodiments, R¹ is phenyl, optionallysubstituted with one or more halogen, CN, C₁-C₄ alkyl, OR⁴, or acombination thereof. In certain embodiments, R¹ is C(═O)—OR^(a), whereinR¹ is C₁-C₄ alkyl.

In some embodiments, R² is H. In some embodiments, R³ is H. In otherembodiments, R² is H and R³ is H.

In some embodiments, in the compound of Formula (II), p is 0. In someembodiments, q is 0. In other embodiments, p is 0 and q is 0.

In some embodiments, in the compound of Formula (II), U is N.

In some embodiments, the compound for use in the invention isrepresented by a compound of Formula (III):

-   -   wherein X, U, R¹, R², R³, m, n, p, and q are defined as anywhere        herein.

In some embodiments, in the compound of Formula (III), X is O, NH, or abond. In some embodiments, X is O. In other embodiments, X is a bond. Incertain embodiments, X is NH.

In some embodiments, in the compound of Formula (III), m and n are each0.

In some embodiments, in the compound of Formula (III), R¹ is aryl orheteroaryl. In some embodiments, R¹ is phenyl, quinolinyl, orisoquinolinyl. In other embodiments, R¹ is phenyl, optionallysubstituted with one or more halogen, CN, C₁-C₄ alkyl, OR⁴, or acombination thereof. In certain embodiments, R¹ is C(═O)—OR^(a), whereinR^(a) is C₁-C₄ alkyl.

In some embodiments, R² is H. In some embodiments, R³ is H. In otherembodiments, R² is H and R³ is H.

In some embodiments, in the compound of Formula (III), p is 0. In someembodiments, q is 0. In other embodiments, p is 0 and q is 0.

In some embodiments, in the compound of Formula (III), U is N.

In some embodiments, the compound for use in the present invention isrepresented by a compound of Formula (IV)

-   -   wherein X, U, R¹, R², R³, m, n, p, and q are defined as anywhere        herein.

In some embodiments, in the compound of Formula (IV), X is O. In otherembodiments, X is S. In certain embodiments, X is NH.

In some embodiments, in the compound of Formula (IV), R¹ is aryl orheteroaryl. In some embodiments, R¹ is phenyl, quinolinyl, orisoquinolinyl. In other embodiments, R¹ is phenyl, optionallysubstituted with one or more halogen, CN, C₁-C₄ alkyl, OR⁴, or acombination thereof. In certain embodiments, R¹ is C(═O)—OR^(a), whereinR^(a) is C₁-C₄ alkyl.

In some embodiments, R² is H. In some embodiments, R³ is H. In otherembodiments, R² is H and R³ is H.

In some embodiments, in the compound of Formula (IV), p is 0. In someembodiments, q is 0. In other embodiments, p is 0 and q is 0.

In some embodiments, in the compound of Formula (IV), U is N.

In some embodiments, the compound for use in the present invention is:

-   -   including salts, hydrates, solvates, polymorphs, optical        isomers, geometrical isomers, enantiomers, diastereomers, and        mixtures thereof.

In some embodiments, the compound for use in the invention isrepresented by a compound of Formula (V):

-   -   wherein X, U, R¹, R², R³, m, n, p, and q are defined as anywhere        herein.

In some embodiments, in the compound of Formula (V), Z is N. In otherembodiments, Z is CH.

In some embodiments, in the compound of Formula (V), R¹ is aryl orheteroaryl. In some embodiments, R¹ is phenyl, quinolinyl, orisoquinolinyl. In other embodiments, R¹ is phenyl, optionallysubstituted with one or more halogen, CN, C₁-C₄ alkyl, OR⁴, or acombination thereof. In certain embodiments, R¹ is C(═O)—OR^(a), whereinR^(a) is C₁-C₄ alkyl. In some embodiments, R² is H. In some embodiments,R³ is H. In other embodiments, R² is H and R³ is H.

In some embodiments, in the compound of Formula (V), p is 0. In someembodiments, q is 0. In other embodiments, p is 0 and q is 0.

In some embodiments, in the compound of Formula (V), U is N.

In some embodiments, the compound for use in the present invention is:

In some embodiments, the compound for use in the present inventioncomprises a di-meta-OMe analog, which in one embodiment is representedby a compound of Formula (24)

In some embodiments, the compound for use in the present invention isrepresented by a compound of Formula (VI):

-   -   wherein U, R¹, R², R³, p, and q are defined as anywhere herein.

In some embodiments, in the compound of Formula (VI), R¹ is aryl orheteroaryl. In some embodiments, R¹ is phenyl, quinolinyl, orisoquinolinyl. In other embodiments, R¹ is phenyl, optionallysubstituted with one or more halogen, CN, C₁-C₄ alkyl, OR⁴, or acombination thereof. In certain embodiments, R¹ is C(═O)—OR^(a), whereinR^(a) is C₁-C₄ alkyl. In some embodiments, R² is H. In some embodiments,R³ is H. In other embodiments, R² is H and R³ is H.

In some embodiments, in the compound of Formula (VI), p is 0. In someembodiments, q is 0. In other embodiments, p is 0 and q is 0.

In some embodiments, in the compound of Formula (VI), U is N.

In some embodiments, the compound for use in the present invention is:

In another embodiment, R¹ is selected from the group consisting ofphenyl, quinolinyl and isoquinolinyl, each of which may independently beunsubstituted or substituted with one or more halogen, OR^(a) orNR^(a)R^(b) wherein R^(a) and R^(b) are each independently selected fromthe group consisting of H or a C₁-C₄ alkyl.

In another embodiment, R¹ is C(═O)—OR^(a) wherein R^(a) is a C₁-C₄alkyl. In other embodiments, R¹ is selected from the group consistingof:

-   -   a) phenyl;    -   b) fluorophenyl;    -   c) difluorophenyl;    -   d) pentafluorophenyl;    -   e) methoxyphenyl;

and

-   -   g) C(═O)—OCH₂CH₃.

Each possibility represents a separate embodiment of the presentinvention.

In some embodiments, in the compound of Formula (II) or in the compoundof Formula (IV), X is S. In one embodiment, when X is S, Z is CH. In analternative embodiment, the following compound is excluded: a compoundof Formula (II) wherein X is S, Z is N, Y is C═O, n is 0, m is 0, R¹ isan unsubstituted or substituted phenyl and p and q are each 0. Inanother embodiment, the following compound is excluded: a compound offormula (II) wherein X is S, Z is N, Y is C═O, n is 0, m is 0, R¹ ispyridinyl and p and q are each 0.

In some embodiments wherein X is S, R¹ is aryl, optionally substitutedwith one or more alkyl, arylalkyl, halogen, NO₂, CN, OR⁴, andNR^(5a)R^(5b), or a combination thereof. In some embodiments wherein Xis S, R¹ is aryl substituted with one or more alkyl, arylalkyl, halogen,NO₂, CN, OR⁴, and NR^(5a)R^(5b), or a combination thereof. In someembodiments wherein X is S, R¹ is aryl substituted with one or morealkyl, arylalkyl, halogen, NO₂, and NR^(5a)R^(5b), or a combinationthereof. In some embodiments wherein X is S, R¹ is phenyl substitutedwith one or more alkyl, arylalkyl, halogen, NO₂, and NR^(5a)R^(5b), or acombination thereof. In some embodiments wherein X is S, R¹ is aryl,optionally substituted with one or more halogen and C₁-C₄ alkyl, or acombination thereof. In some embodiments wherein X is S, R¹ is phenyl,optionally substituted with one or more halogen and C₁-C₄ alkyl, or acombination thereof.

In some embodiments wherein X is S, R¹ is heteroaryl, optionallysubstituted with one or more alkyl, arylalkyl, halogen, NO₂, CN, OR⁴,and NR^(5a)R^(5b), or a combination thereof. In some embodiments whereinX is S, R¹ is not pyridinyl. In some embodiments wherein X is S, R¹ isheteroaryl substituted with one or more alkyl, arylalkyl, halogen, NO₂,CN, OR⁴, and NR^(5a)R^(5b), or a combination thereof. In someembodiments wherein X is S, R¹ is heteroaryl substituted with one ormore alkyl, arylalkyl, halogen, NO₂, and NR^(5a)R^(5b), or a combinationthereof. In some embodiments wherein X is S, R¹ is quinolinyl, orisoquinolinyl. In some embodiments wherein X is S, R¹ is quinolinyl orisoquinolinyl, optionally substituted with one or more alkyl, arylalkyl,halogen, NO₂, CN, OR⁴, and NR^(5a)R^(5b), or a combination thereof. Insome embodiments wherein X is S, R¹ is quinolinyl or isoquinolinyl,optionally substituted with one or more halogen and C₁-C₄ alkyl, or acombination thereof. In certain embodiments wherein X is S, R¹ isC(═O)—OR^(a), wherein R^(a) is C₁-C₄ alkyl.

In one embodiment, the compound represented by the structure of Formula(II) as described herein is as described but having the proviso that:

-   -   (1) when Z is CH, X is NH, S, or a bond;    -   (2) when X is S, Z is CH; and    -   (3) the following compound: a compound of Formula (II) wherein X        is NH, Z is CH, Y is C═O, n is 0, m is 0, R¹ is a phenyl        substituted by one or more alkoxy, and p and q are each 0, is        excluded.

In another embodiment, the compound represented by the structure ofFormula (II) as described herein is as described but having the provisothat:

-   -   (1) when Z is CH, X is NH, S, or a bond and (2) the following        compounds are excluded:    -   (i) a compound of Formula (II) wherein X is NH, Z is CH, Y is        C═O, n is 0, m is 0, R1 is a phenyl substituted by one or more        alkoxy, and p and q are each 0; and    -   (ii) a compound of Formula (II) wherein X is S, Z is N, Y is        C═O, n is 0, m is 0, R1 is an unsubstituted or substituted        phenyl and p and q are each 0.

In another embodiment, the compound represented by the structure ofFormula (II) as described herein is as described but having the provisothat:

-   -   (1) when Z is CH, X is NH, S, or a bond;    -   (2) the following compounds are excluded:        -   (i) a compound of formula (II) wherein X is NH, Z is CH, Y            is C═O, n is 0, m is 0, R¹ is a phenyl substituted by one or            more alkoxy, and p and q are each 0; and        -   (ii) a compound of formula (II) wherein X is S, Z is N, Y is            C═O, n is 0, m is 0, R¹ is pyridinyl and p and q are each 0.

In one embodiment, the compound is represented by Formula (II), whereinn is 0.

According to this aspect and in one embodiment, the compound isrepresented by the structure of Formula (II-a):

In another embodiment, the compound is represented by Formula (II)wherein X is O, Z is N, Y is C═O, n is 0, m is 0, R¹ is phenyl ormethoxyphenyl and p and q are each 0.

In yet another embodiment, the compound is represented by Formula (II)wherein X is S, Z is CH, Y is C═O, n is 0, m is 0, R¹ is phenyl and pand q are each 0.

In one embodiment, the compound is represented by Formula (II) wherein Xis O, Z is N, Y is C═O, n is 0, m is 0, R¹ is methoxyphenyl and p and qare each 0. In another embodiment, the compound is represented byFormula (II), wherein X is O, Z is N, Y is C═O, n is 0, m is 0, R¹ isdifluorophenyl or pentafluorophenyl and p and q are each 0.

In another embodiment the compound is represented by Formula (II),wherein p and q are each 0 (i.e., R² and R³ do not exist).

As demonstrated herein, said compound has unexpectedly been found to bea highly potent and selective cytotoxic agent, exhibiting selectivecytotoxicity towards cancer as well as pre-cancerous cells and benignhyperproliferative disorders, while having little effect on normalcells.

As used herein, in some embodiments, an “alkyl” group refers to anysaturated aliphatic hydrocarbon, including straight-chain andbranched-chain alkyl groups. In one embodiment, the alkyl group has 1-4carbons designated here as C₁-C₄-alkyl. In some embodiments, the alkylgroup has 1-7 carbons designated here as C₁-C₇-alkyl. The alkyl groupmay be unsubstituted or substituted by one or more groups selected fromhalogen, hydroxy, alkoxy carbonyl, amido, alkylamido, dialkylamido,nitro, amino, alkylamino, dialkylamino, carboxyl, thio and thioalkyl.

The term “aryl” used herein alone or as part of another group denotes anaromatic ring system containing from 6-14 ring carbon atoms. The arylring can be a monocyclic, bicyclic, tricyclic and the like. Non-limitingexamples of aryl groups are phenyl, naphthyl including 1-naphthyl and2-naphthyl, and the like. The aryl group can be unsubstituted orsubstituted through available carbon atoms with one or more groupsdefined hereinabove for alkyl.

The term “heteroaryl” used herein alone or as part of another groupdenotes a heteroaromatic system containing at least one heteroatom ringatom selected from nitrogen, sulfur and oxygen. The heteroaryl contains5 or more ring atoms. In some embodiments, the heteroaryl group contains5-10 ring atoms. The heteroaryl group can be monocyclic, bicyclic,tricyclic and the like. Also included in this expression are thebenzoheterocyclic rings. If nitrogen is a ring atom, the presentinvention also contemplates the N-oxides of the nitrogen containingheteroaryls. Nonlimiting examples of heteroaryls include thienyl,benzothienyl, 1-naphthothienyl, thianthrenyl, furyl, benzofuryl,pyrrolyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrimidinyl,pyridazinyl, indolyl, isoindolyl, indazolyl, purinyl, quinolyl (e.g.1-quinolinyl, 2-quinolinyl, 3-quinolinyl, 4-quinolinyl, 5-quinolinyl,6-quinolinyl, 7-quinolinyl and 8-quinolinyl), isoquinolinyl (e.g.,1-isoquinolinyl, 2-isoquinolinyl, 3-isoquinolinyl, 4-isoquinolinyl,5-isoquinolinyl, 6-isoquinolinyl, 7-isoquinolinyl and 8-isoquinolinyl);naphthyridinyl (e.g., 1-naphthyridinyl, 2-naphthyridinyl), quinoxalinyl,quinazolinyl, cinnolinyl, pteridinyl, carbolinyl, thiazolyl, oxazolyl,isothiazolyl, isoxazolyl and the like. The heteroaryl group canoptionally be substituted through available atoms with one or moregroups defined hereinabove for alkyl. The heteroaryl group can beunsubstituted or substituted through available atoms with one or moregroups defined hereinabove for alkyl.

The term “halogen” or “halo” as used herein alone or as part of anothergroup refers to chlorine, bromine, fluorine, and iodine.

The term “hydroxy” refers to an OH group. The terms “alkoxy” refers tothe group OR^(a) wherein R^(a) is a C₁-C₄ alkyl as defined above.Nonlimiting examples of an alkoxy group is methoxy, ethoxy, n-propoxy,isopropoxy, n-butoxy, t-butoxy and like groups.

All stereoisomers of the compounds of the instant invention arecontemplated, either in admixture or in pure or substantially pure form.The compounds of the present invention can have asymmetric centers atany of the atoms. Consequently, the compounds can exist in enantiomericor diastereomeric forms or in mixtures thereof. The present inventioncontemplates the use of any racemates (i.e. mixtures containing equalamounts of each enantiomers), enantiomerically enriched mixtures (i.e.,mixtures enriched for one enantiomer), pure enantiomers ordiastereomers, or any mixtures thereof. The chiral centers can bedesignated as R or S or R,S or d,D, l,L or d,l, D,L. The presentinvention intends to encompass all structural and geometrical isomersincluding cis, trans, E and Z isomers.

One or more of the compounds of the invention, may be present as a salt.The term “salt” encompasses both basic and acid addition salts,including but not limited to, carboxylate salts or salts with aminenitrogens, and include salts formed with the organic and inorganicanions and cations discussed below. Furthermore, the term includes saltsthat form by standard acid-base reactions with basic groups (such asamino groups) and organic or inorganic acids. Such acids includehydrochloric, hydrofluoric, trifluoroacetic, sulfuric, phosphoric,acetic, succinic, citric, lactic, maleic, fumaric, palmitic, cholic,pamoic, mucic, D-glutamic, D-camphoric, glutaric, phthalic, tartaric,lauric, stearic, salicylic, methanesulfonic, benzenesulfonic, sorbic,picric, benzoic, cinnamic, and like acids. Each possibility represents aseparate embodiment of the invention.

The term “organic or inorganic cation” refers to counter-ions for theanion of a salt. The counter-ions are chosen from the alkali andalkaline earth metals, (such as lithium, sodium, potassium, barium,aluminum and calcium); ammonium and mono-, di- and tri-alkyl amines suchas trimethylamine, cyclohexylamine; and the organic cations, such asdibenzylammonium, benzylammonium, 2-hydroxyethylammonium,bis(2-hydroxyethyl)ammonium, phenylethylbenzylammonium, dibenzylethylenediammonium, and like cations. See, for example, “Pharmaceutical Salts,”Berge et al., J. Pharm. Sci., 66:1-19 (1977), which is incorporatedherein by reference.

The present invention also includes the use of solvates of the compoundsof the present invention and salts thereof. “Solvate” means a physicalassociation of a compound of the invention with one or more solventmolecules. This physical association involves varying degrees of ionicand covalent bonding, including hydrogen bonding. In certain instancesthe solvate will be capable of isolation. “Solvate” encompasses bothsolution-phase and isolatable solvates. Non-limiting examples ofsuitable solvates include ethanolates, methanolates and the like.“Hydrate” is a solvate wherein the solvent molecule is water.

The present invention also includes use of polymorphs of the compoundsof the present invention and salts thereof. The term “polymorph” refersto a particular crystalline state of a substance, which can becharacterized by particular physical properties such as X-raydiffraction, IR spectra, melting point, and the like.

Other Compounds

In another embodiment, the compound for use in the present inventioncomprises 2-deoxyglucose (2-DG). In one embodiment, the compoundcomprises a peptide derived from the N-terminus of HK2, which in oneembodiment, is HKVBD. Such peptides derived from the N-terminus of HK2are known in the art and described, for example, in Wolf et al. 2016(Cell. 2016 Jul. 28; 166(3):624-636) and Pastorino et al 2002 (JBC277:7610-8), which are incorporated by reference herein in theirentirety. In another embodiment, the compound comprises a peptidoglycan,which in one embodiment, is a bacterial peptidoglycan. In anotherembodiment, the compound comprises N-acetylglucosamine. In anotherembodiment, the compound comprises glucose-6-phosphate (G6P). In anotherembodiment, the compound comprises Lonidamine. In another embodiment,the compound comprises 3-bromopyruvate. In another embodiment, thecompound comprises clotrimazole. In another embodiment, the compoundcomprises shRNA, which in one embodiment, comprises VDAC shRNA and, inanother embodiment, comprises HK2 shRNA.

Inflammasomes

In one embodiment, the inflammasome is a cellular protein complex thattriggers secretion of one or more cytokines, which, in one embodiment,activates or recruits the immune system.

In one embodiment, the inflammasome is a multiprotein oligomerconsisting of caspase 1, PYCARD, NALP and sometimes caspase 5 (alsoknown as caspase 11 or ICH-3). It is expressed in myeloid cells and is acomponent of the innate immune system.

In one embodiment, the inflammasome as described herein is an NLRPinflammasome.

In one embodiment, the NLRP inflammasome is an NLRP3 inflammasome. Inanother embodiment, the inflammasome is NLRP1 inflammasome. In oneembodiment, the inflammasome is an NLRC inflammasome. In one embodiment,the NLRC inflammasome is an NLRC4 inflammasome.

In one embodiment, the activation of said inflammasome increasesprocessing and/or secretion of interleukin (IL)-1β. In anotherembodiment, the activation of said inflammasome increases processingand/or secretion of IL-18.

In another embodiment, administration of the compounds of the presentinvention increases Caspase-1 activation.

Cancer and Tumors

In one embodiment, the present invention provides a method of treating asolid tumor or hematological malignancy in a subject by inducing animmune response in a subject comprising the step of contacting one ormore cells of said subject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In anotherembodiment, the present invention provides a method of treating a solidtumor or hematological malignancy in a subject by activating an immuneresponse in a subject comprising the step of contacting one or morecells of said subject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In anotherembodiment, the present invention provides a method of treating a solidtumor or hematological malignancy in a subject by potentiating an immuneresponse in a subject comprising the step of contacting one or morecells of said subject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In one embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula (VII) asdescribed herein. In another embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula C, asdescribed herein. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative.

In one embodiment, the present invention provides a method of treating asolid tumor or hematological malignancy in a subject by inducing aninnate immune response in a subject comprising the step of contactingone or more cells of said subject with a therapeutically effectiveamount of an HK2/mitochondria-detaching compound, as described herein.In another embodiment, the present invention provides a method oftreating a solid tumor or hematological malignancy in a subject byactivating an innate immune response in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound, as describedherein. In another embodiment, the present invention provides a methodof treating a solid tumor or hematological malignancy in a subject bypotentiating an innate immune response in a subject comprising the stepof contacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound, as describedherein. In one embodiment, the HK2/mitochondria-detaching compoundcomprises a jasmonic acid derivative. In one embodiment, the jasmonicacid derivative comprises a compound represented by the structure ofFormula (VII) as described herein. In another embodiment, the jasmonicacid derivative comprises a compound represented by the structure ofFormula C, as described herein. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative.

In one embodiment, the present invention provides a method of treating asolid tumor or hematological malignancy in a subject by inducing ahumoral immune response in a subject comprising the step of contactingone or more cells of said subject with a therapeutically effectiveamount of an HK2/mitochondria-detaching compound, as described herein.In another embodiment, the present invention provides a method oftreating a solid tumor or hematological malignancy in a subject byactivating a humoral immune response in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound, as describedherein. In another embodiment, the present invention provides a methodof treating a solid tumor or hematological malignancy in a subject bypotentiating a humoral immune response in a subject comprising the stepof contacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound, as describedherein. In one embodiment, the HK2/mitochondria-detaching compoundcomprises a jasmonic acid derivative. In one embodiment, the jasmonicacid derivative comprises a compound represented by the structure ofFormula (VII) as described herein. In another embodiment, the jasmonicacid derivative comprises a compound represented by the structure ofFormula C, as described herein. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative.

In one embodiment, the present invention provides a method of treating asolid tumor or hematological malignancy in a subject by inducing acell-mediated immune response in a subject comprising the step ofcontacting one or more cells of said subject with a therapeuticallyeffective amount of an HK2/mitochondria-detaching compound, as describedherein. In another embodiment, the present invention provides a methodof treating a solid tumor or hematological malignancy in a subject byactivating a cell-mediated immune response in a subject comprising thestep of contacting one or more cells of said subject with atherapeutically effective amount of an HK2/mitochondria-detachingcompound, as described herein. In another embodiment, the presentinvention provides a method of treating a solid tumor or hematologicalmalignancy in a subject by potentiating a cell-mediated immune responsein a subject comprising the step of contacting one or more cells of saidsubject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In one embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula (VII) asdescribed herein. In another embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula C, asdescribed herein. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative.

In one embodiment, the present invention provides a method of treating asolid tumor or hematological malignancy in a subject by activating Tcells in a subject comprising the step of contacting one or more cellsof said subject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In anotherembodiment, the present invention provides a method of treating a solidtumor or hematological malignancy in a subject by activating NK cells ina subject comprising the step of contacting one or more cells of saidsubject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In anotherembodiment, the present invention provides a method of treating a solidtumor or hematological malignancy in a subject by activating dendriticcells in a subject comprising the step of contacting one or more cellsof said subject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In anotherembodiment, the present invention provides a method of treating a solidtumor or hematological malignancy in a subject by activating mast cellsin a subject comprising the step of contacting one or more cells of saidsubject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In anotherembodiment, the present invention provides a method of treating a solidtumor or hematological malignancy in a subject by activating dendriticcells in a subject comprising the step of contacting one or more cellsof said subject with a therapeutically effective amount of anHK2/mitochondria-detaching compound, as described herein. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In one embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula (VII) asdescribed herein. In another embodiment, the jasmonic acid derivativecomprises a compound represented by the structure of Formula C, asdescribed herein. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative.

In another embodiment, the present invention provides a method oftreating a solid tumor or hematological malignancy in a subject byincreasing inflammation in situ, the method comprising contacting solidtumor or hematological malignancy cells with a therapeutically effectiveamount of an HK2/mitochondria-detaching compound, as described herein.In one embodiment, the HK2/mitochondria-detaching compound comprises ajasmonic acid derivative. In one embodiment, the jasmonic acidderivative comprises a compound represented by the structure of Formula(VII) as described herein. In another embodiment, the jasmonic acidderivative comprises a compound represented by the structure of FormulaC, as described herein. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative.

In still another embodiment, the administration of the compound asdescribed herein reduces growth of the cells of the solid tumor orhematological malignancy by 40%, 50%, 60%, 70%, 80%, 90% or 95% comparedto growth of the cells of the solid tumor or hematological malignancythat have not been treated with the compound.

In one embodiment, the cells contacted with the compound as describedherein are abnormal. In one embodiment, the cells are cancerous.

In one embodiment, a subject as described herein has cancer. In oneembodiment, the term “cancer” in the context of the present inventionincludes all types of neoplasm whether in the form of solid or non-solidtumors, and includes both malignant and premalignant conditions as wellas their metastasis.

In one embodiment, a subject as described herein has a pre-cancerouscondition. In another embodiment, a subject as described herein has abenign hyperproliferative disorder. In another embodiment, said subjecthas cancer.

In one embodiment, the term “pre-cancer” or “pre-malignant” as usedherein interchangeably refers to diseases, syndromes or other conditionsassociated with an increased risk of cancer. Pre-cancer conditions inthe context of the present invention include, but are not limited to:breast calcifications, vaginal intra-epithelial neoplasia, Barrett'sesophagus, atrophic gastritis, dyskeratosis congenital, sideropenicdysphagia, lichen planus, oral sibmucous fibrosis, actinic keratosis,solar elastosis, cervical desplasia, leukoplakia and erythroplakia.

In one embodiment, the term “benign hyperproliferative disorder” as usedherein refers to a condition in which there is an abnormal growth anddifferentiation of cells and an increase in the amount of organic tissuethat results from cell proliferation. The benign hyperproliferativedisorder may be attributed to lack of response or inappropriate responseto regulating factors, or alternatively to dysfunctional regulatingfactors. Non-limiting examples of benign hyperproliferative disorder arepsoriasis and benign prostatic hyperplasia (BPH).

In one embodiment, the cancer is a carcinoma, sarcoma, myeloma,leukemia, or lymphoma. In another embodiment, the cancer is a mixedtype.

In one embodiment, the carcinoma is an adenocarcinoma. In anotherembodiment, the carcinoma is a squamous cell carcinoma.

In one embodiment, the sarcoma comprises osteosarcoma or osteogenicsarcoma (bone); Chondrosarcoma (cartilage); Leiomyosarcoma (smoothmuscle); Rhabdomyosarcoma (skeletal muscle); Mesothelial sarcoma ormesothelioma (membranous lining of body cavities); Fibrosarcoma (fibroustissue); Angiosarcoma or hemangioendothelioma (blood vessels);Liposarcoma (adipose tissue); Glioma or astrocytoma (neurogenicconnective tissue found in the brain); Myxosarcoma (primitive embryonicconnective tissue); and Mesenchymous or mixed mesodermal tumor (mixedconnective tissue types).

In one embodiment, the subject has a myeloma, which, in one embodiment,is cancer that originates in the plasma cells of bone marrow. The plasmacells produce some of the proteins found in blood.

In another embodiment, the subject has a leukemia (“non-solid tumor” or“blood cancer”), which in one embodiment, is a cancer of the bone marrow(the site of blood cell production). In one embodiment, leukemiacomprises myelogenous or granulocytic leukemia (malignancy of themyeloid and granulocytic white blood cell series); Lymphatic,lymphocytic, or lymphoblastic leukemia (malignancy of the lymphoid andlymphocytic blood cell series); and Polycythemia vera or erythremia(malignancy of various blood cell products, but with red cellspredominating).

In another embodiment, the subject has a lymphoma. In one embodiment,lymphomas develop in the glands or nodes of the lymphatic system, anetwork of vessels, nodes, and organs (specifically the spleen, tonsils,and thymus) that purify bodily fluids and produce infection-fightingwhite blood cells, or lymphocytes. Unlike the leukemias, which aresometimes called “non-solid tumors,” lymphomas are “solid cancers.”Lymphomas may also occur in specific organs such as the stomach, breastor brain. These lymphomas are referred to as extranodal lymphomas. Thelymphomas are subclassified into two categories: Hodgkin lymphoma andNon-Hodgkin lymphoma. The presence of Reed-Sternberg cells in Hodgkinlymphoma diagnostically distinguishes Hodgkin lymphoma from Non-Hodgkinlymphoma.

Mixed Type cancers contain several types of cells. The type componentsmay be within one category or from different categories. Some examplesare: adenosquamous carcinoma; mixed mesodermal tumor; carcinosarcoma;teratocarcinoma

As used herein, the term “cancer” includes the above categories ofcarcinoma, sarcoma, myeloma, leukemia, lymphoma and mixed type tumors.In particular, the term cancer includes: lymphoproliferative disorders,breast cancer, ovarian cancer, prostate cancer, cervical cancer,endometrial cancer, lung cancer, bone cancer, liver cancer, stomachcancer, bladder cancer, colon cancer, colorectal cancer, pancreaticcancer, cancer of the thyroid, head and neck cancer, cancer of thecentral nervous system, brain cancer, cancer of the peripheral nervoussystem, skin cancer, kidney cancer, as well as metastases of all theabove. More particularly, as used herein the term may refer to:hepatocellular carcinoma, hematoma, hepatoblastoma, rhabdomyosarcoma,esophageal carcinoma, thyroid carcinoma, ganglioblastoma, fibrosarcoma,myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma,angiosarcoma, endotheliosarcoma, Ewing's tumor, leimyosarcoma,rhabdotheliosarcoma, invasive ductal carcinoma, papillaryadenocarcinoma, melanoma, squamous cell carcinoma, basal cell carcinoma,adenocarcinoma (well differentiated, moderately differentiated, poorlydifferentiated or undifferentiated), renal cell carcinoma,hypernephroma, hypernephroid adenocarcinoma, bile duct carcinoma,choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor, testiculartumor, lung carcinoma including small cell, non-small and large celllung carcinoma, bladder carcinoma, glioma, astrocyoma, medulloblastoma,craniopharyngioma, ependymoma, pinealoma, retinoblastoma, neuroblastoma,colon carcinoma, rectal carcinoma, hematopoietic malignancies includingall types of leukemia and lymphoma including: acute myelogenousleukemia, acute myelocytic leukemia, acute lymphocytic leukemia, chronicmyelogenous leukemia, chronic lymphocytic leukemia, mast cell leukemia,multiple myeloma, myeloid lymphoma, Hodgkin's lymphoma, non-Hodgkin'slymphoma.

In one embodiment, a subject as described herein has been diagnosed witha lymphoma. In one embodiment, the lymphoma is a cutaneous T-celllymphoma (CTCL). In one embodiment, CTCL occurs when T cells becomecancerous. In one embodiment, CTCL affect the skin, causing differenttypes of skin lesions.

In one embodiment, the CTCL is Mycosis fungoides. In one embodiment,Mycosis fungoides or Alibert-Bazin syndrome or granuloma fungoides, isthe most common form of cutaneous T-cell lymphoma. In one embodiment,Mycosis fungoides generally affects the skin, but may progressinternally over time. In another embodiment, the CTCL is Sézarysyndrome. In one embodiment, Sézary syndrome is an aggressive form ofCTCL.

Thus, in one embodiment, the cancer is a skin tumor.

In one embodiment, the skin tumor is a benign hyperproliferative skinlesion, a pre-cancerous tumor or a cancerous tumor.

In one embodiment, the benign hyperproliferative skin lesion ispsoriasis. In another embodiment, the benign hyperproliferative skinlesion is Ledderhose Disease, Dupuytren's Disease, a keloid or ahypertrophic scar.

In one embodiment, the pre-cancerous tumor is actinic keratosis. In afurther embodiment, the pre-cancerous tumor is Bowen's Disease.

In one embodiment, the cancerous tumor is a melanoma. In anotherembodiment, the cancerous tumor is a basal cell carcinoma, a squamouscell carcinoma or a cutaneous T-cell lymphoma.

In one embodiment, the cutaneous T-cell lymphoma is mycosis fungoides orSezary syndrome.

In one embodiment, the cancer comprises a solid tumor.

In one embodiment, the solid tumor comprises stomach cancer, coloncancer, kidney cancer, pancreatic cancer, thyroid cancer, head and neckcancer, or a combination thereof.

In another embodiment, the cancer comprises prostate cancer. In oneembodiment, prostate cancer comprises castration resistant prostatecancer (CRPC), which, in one embodiment, comprises metastatic castrationresistant prostate cancer (mCRPC).

In another embodiment, the cancer comprises breast cancer. In oneembodiment, breast cancer comprises triple negative breast cancer(TNBC).

In one embodiment, the solid tumor comprises lung cancer.

In one embodiment, the lung cancer comprises small cell lung carcinoma,non-small cell lung carcinoma, or large cell lung carcinoma.

In one embodiment, the solid tumor comprises melanoma.

In one embodiment, the cancer comprises squamous cell carcinoma (SCC),basal cell carcinoma, or a combination thereof.

In another embodiment, the cancer is an adenocarcinoma of the stomach orgastroesophageal junction, Dermatofibrosarcoma protuberans,Endocrine/neuroendocrine tumors, Gastrointestinal stromal tumor, Giantcell tumor of the bone, Kaposi sarcoma,Myelodysplastic/myeloproliferative disorders, Ovarianepithelial/fallopian tube/primary peritoneal cancers, Soft tissuesarcoma, Systemic mastocytosis, Germ cell tumor, or a combinationthereof.

In one embodiment, the cancer comprises a hematological malignancy.

In one embodiment, the hematological malignancy comprises leukemia.

In one embodiment, the leukemia comprises lymphoblastic leukemia, which,in one embodiment comprises acute lymphoblastic leukemia (ALL), acutemyeloid leukemia (AML), chronic lymphocytic leukemia (CLL), and chronicmyeloid leukemia (CML).

In another embodiment, the hematological malignancy comprises alymphoma, which in one embodiment is Non-Hodgkin lymphoma.

Additional Therapeutic Agents

In another embodiment, any of the methods of the present inventionfurther comprise the step of contacting one or more cells of saidsubject with a therapeutic agent, which in one embodiment, comprises ananti-cancer drug.

In one embodiment, the anti-cancer drug comprises a chemotherapeuticagent.

In one embodiment, the chemotherapeutic agent comprises Imiquimod,5-Fluorouracil (5-FU), diclofenac, ingenol mebutate, or a combinationthereof.

In another embodiment, the chemotherapeutic agent comprises Bleomycin,capecitabine, cisplatin, Cyclophosphamide, dacarbazine, Doxorubicin,Epirubicin, etoposide, folinic acid, Methotrexate, Mustine, oxaliplatin,prednisolone, procarbazine, vinblastine, vincristine, or a combinationthereof.

In another embodiment, the provided methods further comprise removal ofthe skin tumor by Mohs surgery.

Targeted Therapies

In another embodiment, the present invention provides a method asdescribed herein comprising contacting one or more cells of said subjectwith a compound as described herein and a targeted therapy.

In one embodiment, the present invention provides methods of treatingcancer comprising administering a compound as described herein incombination with one or more targeted therapies.

In one embodiment, the immunotherapeutic compound is targeted toparticular molecules expressed abnormally by cancer cells. In oneembodiment, the targeted therapy comprises a hormone therapy, signaltransduction inhibitor, gene expression modulator, apoptosis inducer,angiogenesis inhibitor, immunotherapy, or toxin delivery molecules.

In one embodiment, the targeted therapy utilizes small molecules. Inanother embodiment, the targeted therapy utilizes antibodies, which, inone embodiment, are monoclonal antibodies.

In one embodiment, the immunotherapeutic compound comprises abirateroneacetate (Zytiga®), ado-trastuzumab emtansine (Kadcyla®), afatinibdimaleate (Gilotrif®), alectinib (Alecensa®), alemtuzumab (Campath®),Alitretinoin (Panretin®), anastrozole (Arimidex®), Atezolizumab(Tecentriq™), axitinib (Inlyta®), belinostat (Beleodaq®), Bevacizumab(Avastin®), bexarotene (Targretin®), blinatumomab (Blincyto®),bortezomib (Velcade®), bosutinib (Bosulif®), brentuximab vedotin(Adcetris®), Cabazitaxel (Jevtana®), cabozantinib (Cabometyx™),Cabozantinib (Cometriq®), carfilzomib (Kyprolis®), ceritinib(LDK378/Zykadia™), Cetuximab (Erbitux®), cobimetinib (Cotellic™),crizotinib (Xalkori®), dabrafenib (Tafinlar®), daratumumab (Darzalex™),dasatinib (Sprycel®), denileukin diftitox (Ontak®), Denosumab (Xgeva®),Dinutuximab (Unituxin™), elotuzumab (Empliciti™), enzalutamide(Xtandi®), Erlotinib (Tarceva®), everolimus (Afinitor®), exemestane(Aromasin®), fulvestrant (Faslodex®), gefitinib (Iressa®), Ibritumomabtiuxetan (Zevalin®), ibrutinib (Imbruvica®), idelalisib (Zydelig®),Imatinib mesylate (Gleevec®), Ipilimumab (Yervoy®), ixazomib citrate(Ninlaro®), Lanreotide acetate (Somatuline® Depot), lapatinib (Tykerb®),lenvatinib mesylate (Lenvima®), letrozole (Femara®), necitumumab(Portrazza™), nilotinib (Tasigna®), nivolumab (Opdivo®), obinutuzumab(Gazyva®), ofatumumab (Arzerra®), olaparib (Lynparza™), olaratumab(Lartruvo™), osimertinib (Tagrisso™), palbociclib (Ibrance®),panitumumab (Vectibix®), panobinostat (Farydak®), pazopanib (Votrient®),pembrolizumab (Keytruda®), pertuzumab (Perjeta®), pralatrexate(Folotyn®), radium 223 dichloride (Xofigo®), ramucirumab (Cyramza®),regorafenib (Stivarga®), rituximab (Rituxan®), romidepsin (Istodax®),ruxolitinib phosphate (Jakafi®), siltuximab (Sylvant®), sonidegib(Odomzo®), sorafenib (Nexavar®), sunitinib (Sutent®), tamoxifen(Nolvadex), temsirolimus (Torisel®), toremifene (Fareston®), trametinib(Mekinist®), Trastuzumab (Herceptin®), Tretinoin (Vesanoid®), vandetanib(Caprelsa®), vemurafenib (Zelboraf®), venetoclax (Venclexta™),Vismodegib (Erivedge®), vorinostat (Zolinza®), ziv-aflibercept(Zaltrap®), or a combination thereof.

In another embodiment, methods of the present invention further comprisethe step of contacting one or more cells of the subject with animmunotherapeutic compound.

Immunotherapeutic Compounds

In one embodiment, an immunotherapy as described herein is a monoclonalantibody that recognizes specific molecules on the surface of cancercells. In one embodiment, binding of the monoclonal antibody to thetarget molecule results in the immune destruction of cells that expressthat target molecule. In another embodiment, the antibody binds tocertain immune cells to enhance their actions on cancer cells.

In one embodiment, the immunotherapeutic compound comprises imatinib ortrastuzumab.

In one embodiment, the immunotherapeutic compound comprises a checkpointinhibitor.

In one embodiment, the checkpoint inhibitor comprises a Programmed cellDeath protein 1 (PD1) inhibitor or a Programmed cell Death Ligand 1(PD-L1) inhibitor.

In one embodiment, the PD-1 or PD-L1 inhibitor is an antibody.

In one embodiment, the antibody comprises Nivolumab, Pembrolizumab,Pidilizumab, Avelumab, BMS 936559, or MPDL328OA.

Chimeric Antigen Receptor-Expressing T-Cells (CAR T-Cells)

In one embodiment, the immunotherapeutic compound comprises chimericantigen receptor T cells (CAR T-cells).

In another embodiment, an immunotherapy as described herein is adoptivecell transfer (ACT) therapy. In one embodiment, ACT therapy comprisescytotoxic T-cells from a patient or donor that are engineered to expressa chimeric antigen receptor (CAR T-cells) targeted to a tumor specificantigen expressed on the surface of the tumor cells. These CAR T-cellsare then cytotoxic only to cells expressing the tumor specific antigen.

In one embodiment, chimeric antigen receptors (CARs) are a type ofantigen-targeted receptor composed of intracellular T-cell signalingdomains fused to extracellular tumor-binding moieties, most commonlysingle-chain variable fragments (scFvs) from monoclonal antibodies. CARsdirectly recognize cell surface antigens, independent of MHC-mediatedpresentation, permitting the use of a single receptor construct specificfor any given antigen in all patients. Initial CARs fusedantigen-recognition domains to the CD3 activation chain of the T-cellreceptor (TCR) complex. While these first generation CARs induced T-celleffector function in vitro, they were largely limited by poor antitumorefficacy in vivo. Subsequent CAR iterations have included secondarycostimulatory signals in tandem with CD3ζ, including intracellulardomains from CD28 or a variety of TNF receptor family molecules such as4-1BB (CD137) and OX40 (CD134). Further, third generation receptorsinclude two costimulatory signals in addition to CD3ζ, most commonlyfrom CD28 and 4-1BB. Second and third generation CARs dramaticallyimproved antitumor efficacy, in some cases inducing complete remissionsin patients with advanced cancer.

In one embodiment, a CAR T-cell is an immunoresponsive cell comprisingan antigen receptor, which is activated when its receptor binds to itsantigen.

In one embodiment, the CAR T-cells used in the compositions and methodsas disclosed herein are first generation CAR T-cells. In anotherembodiment, the CAR T-cells used in the compositions and methods asdisclosed herein are second generation CAR T-cells. In anotherembodiment, the CAR T-cells used in the compositions and methods asdisclosed herein are third generation CAR T-cells. In anotherembodiment, the CAR T-cells used in the compositions and methods asdisclosed herein are fourth generation CAR T-cells.

In one embodiment, first-generation CARs have one signaling domain,typically the cytoplasmic signaling domain of the CD3 TCRζ chain.

In another embodiment, the CAR T-cells as disclosed herein are secondgeneration CAR T-cells. In another embodiment, CAR T-cells as disclosedherein comprise a tripartite chimeric receptor (TPCR). In oneembodiment, CAR T-cells as disclosed herein, comprise one or moresignaling moieties that activate naïve T-cells in a co-stimulationindependent manner. In another embodiment, the CAR T-cells furtherencode one or more members of the tumor necrosis factor receptor family,which in one embodiment, is CD27, 4-1BB (CD137), or OX40 (CD134), or acombination thereof.

Third-generation CAR T-cells attempt to harness the signaling potentialof 2 costimulatory domains: in one embodiment, the CD28 domain followedby either the 4-1BB or OX-40 signaling domains. In another embodiment,the CAR T-cells used in the compositions and methods as disclosed hereinfurther encode a co-stimulatory signaling domain, which in oneembodiment is CD28. In another embodiment, the signaling domain is theCD3ζ-chain, CD97, GDI 1a-CD18, CD2, ICOS, CD27, CD154, CDS, OX40, 4-1BB,CD28 signaling domain, or combinations thereof.

In one embodiment, telomere length and replicative capacity correlatewith the engraftment efficiency and antitumor efficacy of adoptivelytransferred T-cell lines. In one embodiment, CD28 stimulation maintainstelomere length in T-cells.

In one embodiment, CAR-modified T-cell potency may be further enhancedthrough the introduction of additional genes, including those encodingproliferative cytokines (ie, IL-12) or costimulatory ligands (ie,4-1BBL), thus producing “armored” fourth-generation CAR-modifiedT-cells. In one embodiment, “armored CAR T-cells,” are CAR T-cells whichare protected from the inhibitory tumor microenvironment. In anotherembodiment, the “armored” CAR technology incorporates the localsecretion of soluble signaling proteins to amplify the immune responsewithin the tumor microenvironment with the goal of minimizing systemicside effects. In one embodiment, the signaling protein signal is IL-12,which can stimulate T-cell activation and recruitment. In oneembodiment, “armored” CAR technology is especially useful in solid tumorindications, in which microenvironment and potent immunosuppressivemechanisms have the potential to make the establishment of a robustanti-tumor response more challenging.

In one embodiment, CAR T-cells are genetically modified to encodemolecules involved in the prevention of apoptosis, the remodeling of thetumor microenvironment, induction of homeostatic proliferation, andchemokine receptors that promote directed T-cell homing.

In one embodiment, the CAR binds to an epitope of an antigen via anantibody or an antibody fragment that is directed to the antigen. Inanother embodiment, the antibody is a monoclonal antibody. In anotherembodiment, the antibody is a polyclonal antibody. In anotherembodiment, the antibody fragment is a single-chain variable fragment(scFv).

In another embodiment, the CAR T-cells of the compositions as disclosedherein bind to a tumor associated antigen (TAA). In another embodiment,said tumor associated antigen is: Mucin 1, cell surface associated(MUC1) or polymorphic epithelial mucin (PEM), Arginine-rich, mutated inearly stage tumors (Armet), Heat Shock Protein 60 (HSP60), calnexin(CANX), methylenetetrahydrofolate dehydrogenase (NADP+ dependent) 2,methenyltetrahydrofolate cyclohydrolase (MTHFD2), fibroblast activationprotein (FAP), matrix metallopeptidase (MMP6), B Melanoma Antigen-1(BAGE-1), aberrant transcript of N-acetyl glucosaminyl transferase V(GnTV), Q5H943, Carcinoembryonic antigen (CEA), Pmel, Kallikrein-4,Mammaglobin-1, MART-1, GPR143-OA1, prostate specific antigen (PSA),TRP1, Tyrosinase, FGP-5, NEU proto-oncogene, Aft, MMP-2, prostatespecific membrane antigen (PSMA), Telomerase-associated protein-2,Prostatic acid phosphatase (PAP), Uroplakin II or Proteinase 3.

In another embodiment, the CAR binds to CD19 or CD20 to target B cellsin the case where one would like to destroy B cells as in leukemia. Inanother embodiment, the CAR binds to ROR1, CD22, or GD2. In anotherembodiment, the CAR binds to NY-ESO-1. In another embodiment, the CARbinds to MAGE family proteins. In another embodiment, the CAR binds tomesothelin. In another embodiment, the CAR binds to c-erbB2. In anotherembodiment, the CAR binds to mutational antigens that are tumorspecific, such as BRAFV600E mutations and BCR-ABL translocations. Inanother embodiment, the CAR binds to viral antigens which aretumor-specific, such as EBV in HD, HPV in cervical cancer, andpolyomavirus in Merkel cancer.

In another embodiment, the CAR T-cell binds to Her2/neu. In anotherembodiment, the CAR T-cell binds to EGFRvIII.

In one embodiment, the chimeric antigen receptor (CAR) T-cell binds theCD19 antigen. In another embodiment, the CAR binds the CD22 antigen. Inanother embodiment, the CAR binds to alpha folate receptor. In anotherembodiment, the CAR binds to CAIX. In another embodiment, the CAR bindsto CD20. In another embodiment, the CAR binds to CD23. In anotherembodiment, the CAR binds to CD24. In another embodiment, the CAR bindsto CD30. In another embodiment, the CAR binds to CD33. In anotherembodiment, the CAR binds to CD38. In another embodiment, the CAR bindsto CD44v6. In another embodiment, the CAR binds to CD44v7/8.

In another embodiment, the CAR binds to CD123. In another embodiment,the CAR binds to CD171. In another embodiment, the CAR binds tocarcinoembryonic antigen (CEA). In another embodiment, the CAR binds toEGFRvIII. In another embodiment, the CAR binds to EGP-2. In anotherembodiment, the CAR binds to EGP-40. In another embodiment, the CARbinds to EphA2. In another embodiment, the CAR binds to Erb-B2. Inanother embodiment, the CAR binds to Erb-B 2,3,4. In another embodiment,the CAR binds to Erb-B3/4. In another embodiment, the CAR binds to FBP.In another embodiment, the CAR binds to fetal acetylcholine receptor. Inanother embodiment, the CAR binds to GD₂. In another embodiment, the CARbinds to G_(D3). In another embodiment, the CAR binds to HER2. Inanother embodiment, the CAR binds to HMW-MAA. In another embodiment, theCAR binds to IL-11Ralpha. In another embodiment, the CAR binds toIL-13Ralpha1. In another embodiment, the CAR binds to KDR. In anotherembodiment, the CAR binds to kappa-light chain. In another embodiment,the CAR binds to Lewis Y. In another embodiment, the CAR binds toL1-cell adhesion molecule. In another embodiment, the CAR binds toMAGE-A1. In another embodiment, the CAR binds to mesothelin. In anotherembodiment, the CAR binds to CMV infected cells. In another embodiment,the CAR binds to MUC1. In another embodiment, the CAR binds to MUC16. Inanother embodiment, the CAR binds to NKG2D ligands. In anotherembodiment, the CAR binds to NY-ESO-1 (amino acids 157-165). In anotherembodiment, the CAR binds to oncofetal antigen (h5T4). In anotherembodiment, the CAR binds to PSCA. In another embodiment, the CAR bindsto PSMA. In another embodiment, the CAR binds to ROR1. In anotherembodiment, the CAR binds to TAG-72. In another embodiment, the CARbinds to VEGF-R2 or other VEGF receptors. In another embodiment, the CARbinds to B7-H6. In another embodiment, the CAR binds to CA9. In anotherembodiment, the CAR binds to avif integrin. In another embodiment, theCAR binds to 8H9. In another embodiment, the CAR binds to NCAM. Inanother embodiment, the CAR binds to fetal acetylcholine receptor.

In another embodiment, the chimeric antigen receptor (CAR) T-celltargets the CD19 antigen, and has a therapeutic effect on subjects withB-cell malignancies, ALL, Follicular lymphoma, CLL, and Lymphoma. Inanother embodiment, the CAR T-cell targets the CD22 antigen, and has atherapeutic effect on subjects with B-cell malignancies. In anotherembodiment, the CAR T-cell targets alpha folate receptor or folatereceptor alpha, and has a therapeutic effect on subjects with ovariancancer or epithelial cancer. In another embodiment, the CAR T-celltargets CAIX or G250/CAIX, and has a therapeutic effect on subjects withrenal cell carcinoma. In another embodiment, the CAR T-cell targetsCD20, and has a therapeutic effect on subjects with Lymphomas, B-cellmalignancies, B-cell lymphomas, Mantle cell lymphoma and, indolentB-cell lymphomas. In another embodiment, the CAR T-cell targets CD23,and has a therapeutic effect on subjects with CLL. In anotherembodiment, the CAR T-cell targets CD24, and has a therapeutic effect onsubjects with pancreatic adenocarcinoma. In another embodiment, the CART-cell targets CD30, and has a therapeutic effect on subjects withLymphomas or Hodgkin lymphoma. In another embodiment, the CAR T-celltargets CD33, and has a therapeutic effect on subjects with AML. Inanother embodiment, the CAR T-cell targets CD38, and has a therapeuticeffect on subjects with Non-Hodgkin lymphoma. In another embodiment, theCAR T-cell targets CD44v6, and has a therapeutic effect on subjects withseveral malignancies. In another embodiment, the CAR T-cell targetsCD44v7/8, and has a therapeutic effect on subjects with cervicalcarcinoma. In another embodiment, the CAR T-cell targets CD123, and hasa therapeutic effect on subjects with myeloid malignancies. In anotherembodiment, the CAR T-cell targets CEA, and has a therapeutic effect onsubjects with colorectal cancer. In another embodiment, the CAR T-celltargets EGFRvIII, and has a therapeutic effect on subjects withGlioblastoma. In another embodiment, the CAR T-cell targets EGP-2, andhas a therapeutic effect on subjects with multiple malignancies. Inanother embodiment, the CAR T-cell targets EGP-40, and has a therapeuticeffect on subjects with colorectal cancer. In another embodiment, theCAR T-cell targets EphA2, and has a therapeutic effect on subjects withGlioblastoma. In another embodiment, the CAR T-cell targets Erb-B2 orErbB3/4, and has a therapeutic effect on subjects with Breast cancer andothers, prostate cancer, colon cancer, various tumors. In anotherembodiment, the CAR T-cell targets Erb-B 2,3,4, and has a therapeuticeffect on subjects with Breast cancer and others. In another embodiment,the CAR T-cell targets FBP, and has a therapeutic effect on subjectswith Ovarian cancer. In another embodiment, the CAR T-cell targets fetalacetylcholine receptor, and has a therapeutic effect on subjects withRhabdomyosarcoma. In another embodiment, the CAR T-cell targets G_(D2),and has a therapeutic effect on subjects with Neuroblastoma, melanoma,or Ewing's sarcoma. In another embodiment, the CAR T-cell targets GD₃,and has a therapeutic effect on subjects with Melanoma. In anotherembodiment, the CAR T-cell targets HER2, and has a therapeutic effect onsubjects with medulloblastoma, pancreatic adenocarcinoma, Glioblastoma,Osteosarcoma, or Ovarian cancer. In another embodiment, the CAR T-celltargets HMW-MAA, and has a therapeutic effect on subjects with Melanoma.In another embodiment, the CAR T-cell targets IL-11 Ralpha, and has atherapeutic effect on subjects with Osteosarcoma. In another embodiment,the CAR T-cell targets IL-13Ralpha1, and has a therapeutic effect onsubjects with Glioma, Glioblastoma, or medulloblastoma. In anotherembodiment, the CAR T-cell targets IL-13 receptor alpha2, and has atherapeutic effect on subjects with several malignancies. In anotherembodiment, the CAR T-cell targets KDR, and has a therapeutic effect onsubjects with tumors by targeting tumor neovasculature. In anotherembodiment, the CAR T-cell targets kappa-light chain, and has atherapeutic effect on subjects with B-cell malignancies (B-NHL, CLL). Inanother embodiment, the CAR T-cell targets Lewis Y, and has atherapeutic effect on subjects with various carcinomas orepithelial-derived tumors.

In another embodiment, the CAR T-cell targets L1-cell adhesion molecule,and has a therapeutic effect on subjects with Neuroblastoma. In anotherembodiment, the CAR T-cell targets MAGE-A1 or HLA-A MAGE A1, and has atherapeutic effect on subjects with Melanoma. In another embodiment, theCAR T-cell targets mesothelin, and has a therapeutic effect on subjectswith Mesothelioma. In another embodiment, the CAR T-cell targets CMVinfected cells, and has a therapeutic effect on subjects with CMV. Inanother embodiment, the CAR T-cell targets MUC1, and has a therapeuticeffect on subjects with breast or ovarian cancer. In another embodiment,the CAR T-cell targets MUC16, and has a therapeutic effect on subjectswith ovarian cancer. In another embodiment, the CAR T-cell targets NKG2Dligands, and has a therapeutic effect on subjects with myeloma, ovarian,and other tumors. In another embodiment, the CAR T-cell targets NY-ESO-1(157-165) or HLA-A2 NY-ESO-1, and has a therapeutic effect on subjectswith multiple myeloma. In another embodiment, the CAR T-cell targetsoncofetal antigen (h5T4), and has a therapeutic effect on subjects withvarious tumors. In another embodiment, the CAR T-cell targets PSCA, andhas a therapeutic effect on subjects with prostate carcinoma. In anotherembodiment, the CAR T-cell targets PSMA, and has a therapeutic effect onsubjects with prostate cancer/tumor vasculature. In another embodiment,the CAR T-cell targets ROR1, and has a therapeutic effect on subjectswith B-CLL and mantle cell lymphoma. In another embodiment, the CART-cell targets TAG-72, and has a therapeutic effect on subjects withadenocarcinomas or gastrointestinal cancers. In another embodiment, theCAR T-cell targets VEGF-R2 or other VEGF receptors, and has atherapeutic effect on subjects with tumors by targeting tumorneovasculature. In another embodiment, the CAR T-cell targets CA9, andhas a therapeutic effect on subjects with renal cell carcinoma. Inanother embodiment, the CAR T-cell targets CD171, and has a therapeuticeffect on subjects with renal neuroblastoma. In another embodiment, theCAR T-cell targets NCAM, and has a therapeutic effect on subjects withneuroblastoma. In another embodiment, the CAR T-cell targets fetalacetylcholine receptor, and has a therapeutic effect on subjects withrhabdomyosarcoma. In another embodiment, the CAR binds to one of thetarget antigens listed in Table 1 of Sadelain et al. (Cancer Discov.2013 April; 3(4): 388-398), which is incorporated by reference herein inits entirety. In another embodiment, CAR T-cells bind to carbohydrate orglycolipid structures.

In one embodiment the CAR binds to an angiogenic factor, therebytargeting tumor vasculature. In one embodiment, the angiogenic factor isVEGFR2. in another embodiment, the angiogenic factor is endoglin. Inanother embodiment, an angiogenic factor disclosed herein is Angiogenin;Angiopoietin-1; Del-1; Fibroblast growth factors: acidic (aFGF) andbasic (bFGF); Follistatin; Granulocyte colony-stimulating factor(G-CSF); Hepatocyte growth factor (HGF)/scatter factor (SF);Interleukin-8 (IL-8); Leptin; Midkine; Placental growth factor;Platelet-derived endothelial cell growth factor (PD-ECGF);Platelet-derived growth factor-BB (PDGF-BB); Pleiotrophin (PTN);Progranulin; Proliferin; Transforming growth factor-alpha (TGF-alpha);Transforming growth factor-beta (TGF-beta); Tumor necrosis factor-alpha(TNF-alpha); Vascular endothelial growth factor (VEGF)/vascularpermeability factor (VPF). In another embodiment, an angiogenic factoris an angiogenic protein. In one embodiment, a growth factor is anangiogenic protein. In one embodiment, an angiogenic protein for use inthe compositions and methods disclosed herein is Fibroblast growthfactors (FGF); VEGF; VEGFR and Neuropilin 1 (NRP-1); Angiopoietin 1(Ang1) and Tie2; Platelet-derived growth factor (PDGF; BB-homodimer) andPDGFR; Transforming growth factor-beta (TGF-β), endoglin and TGF-βreceptors; monocyte chemotactic protein-1 (MCP-1); Integrins αVβ3, αVβ5and α5β1; VE-cadherin and CD31; ephrin; plasminogen activators;plasminogen activator inhibitor-1; Nitric oxide synthase (NOS) andCOX-2; AC 133; or Id1/Id3. In one embodiment, an angiogenic protein foruse in the compositions and methods disclosed herein is an angiopoietin,which in one embodiment, is Angiopoietin 1, Angiopoietin 3, Angiopoietin4 or Angiopoietin 6. In one embodiment, endoglin is also known as CD105;EDG; HHT1; ORW; or ORW1. In one embodiment, endoglin is a TGFbetaco-receptor.

In another embodiment, the CAR T-cells bind to an antigen associatedwith an infectious agent. In one embodiment, the infectious agent isMycobacterium tuberculosis. In one embodiment, said Mycobacteriumtuberculosis associated antigen is: Antigen 85B, Lipoprotein IpqH, ATPdependent helicase putative, uncharacterized protein Rv0476/MTO4941precursor or uncharacterized protein Rv1334/MT1376 precursor.

In another embodiment, the CAR binds to an antibody. In one embodiment,the CAR T-cell is an “antibody-coupled T-cell receptor” (ACTR).According to this embodiment, the CAR T-cell is a universal CAR T-cell.In another embodiment, the CAR T-cell having an antibody receptor isadministered before, after, or at the same time as the antibody isadministered and then binds to the antibody, bringing the T-cell inclose proximity to the tumor or cancer. In another embodiment, theantibody is directed against a tumor cell antigen. In anotherembodiment, the antibody is directed against CD20. In anotherembodiment, the antibody is rituximab.

In another embodiment, the antibody is Trastuzumab (Herceptin;Genentech): humanized IgG1, which is directed against ERBB2. In anotherembodiment, the antibody is Bevacizumab (Avastin; Genentech/Roche):humanized IgG1, which is directed against VEGF. In another embodiment,the antibody is Cetuximab (Erbitux; Bristol-Myers Squibb): chimerichuman-murine IgG1, which is directed against EGFR. In anotherembodiment, the antibody is Panitumumab (Vectibix; Amgen): human IgG2,which is directed against EGFR. In another embodiment, the antibody isIpilimumab (Yervoy; Bristol-Myers Squibb): IgG1, which is directedagainst CTLA4.

In another embodiment, the antibody is Alemtuzumab (Campath; Genzyme):humanized IgG1, which is directed against CD52. In another embodiment,the antibody is Ofatumumab (Arzerra; Genmab): human IgG1, which isdirected against CD20. In another embodiment, the antibody is Gemtuzumabozogamicin (Mylotarg; Wyeth): humanized IgG4, which is directed againstCD33. In another embodiment, the antibody is Brentuximab vedotin(Adcetris; Seattle Genetics): chimeric IgG1, which is directed againstCD30. In another embodiment, the antibody is 90Y-labelled ibritumomabtiuxetan (Zevalin; IDEC Pharmaceuticals): murine IgG1, which is directedagainst CD20. In another embodiment, the antibody is 1311-labelledtositumomab (Bexxar; GlaxoSmithKline): murine IgG2, which is directedagainst CD20.

In another embodiment, the antibody is Ramucirumab, which is directedagainst vascular endothelial growth factor receptor-2 (VEGFR-2). Inanother embodiment, the antibody is ramucirumab (Cyramza Injection, EliLilly and Company), blinatumomab (BLINCYTO, Amgen Inc.), pembrolizumab(KEYTRUDA, Merck Sharp & Dohme Corp.), obinutuzumab (GAZYVA, Genentech,Inc.; previously known as GA101), pertuzumab injection (PERJETA,Genentech, Inc.), or denosumab (Xgeva, Amgen Inc.). In anotherembodiment, the antibody is Basiliximab (Simulect; Novartis). In anotherembodiment, the antibody is Daclizumab (Zenapax; Roche).

In another embodiment, the antibody to which the CAR T-cell is coupledis directed to a tumor or cancer antigen or a portion thereof, that isdescribed herein and/or that is known in the art. In another embodiment,the antibody to which the CAR T-cell is couples is directed to atumor-associated antigen. In another embodiment, the antibody to whichthe CAR T-cell is couples is directed to a tumor-associated antigen or aportion thereof that is an angiogenic factor.

In another embodiment, the antibody to which the CAR T-cell is coupledis directed to a tumor or cancer antigen or a portion thereof, that isdescribed herein and/or that is known in the art.

Infections

In another embodiment, a subject as described herein suffers from aninfection.

In one embodiment, the present invention provides a method ofsuppressing, inhibiting, decreasing the risk of, or preventing aninfection in a subject comprising contacting one or more cells of saidsubject with a therapeutically effective amount of a compound thatpromotes the detachment of HK2 from mitochondria.

In one embodiment, the HK2/mitochondria-detaching compound comprises ajasmonic acid derivative. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative.In one embodiment, the compound is represented by the structure ofFormula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof. In another embodiment,the compound is represented by the structure of Formula C, as describedherein.

In one embodiment, the present invention provides a method of treatingan infection in a subject comprising contacting one or more cells ofsaid subject with a therapeutically effective amount of a compound thatpromotes the detachment of HK2 from mitochondria. In one embodiment, theHK2/mitochondria-detaching compound comprises a jasmonic acidderivative. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative. In one embodiment, thecompound is represented by the structure of Formula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof. In another embodiment,the compound is represented by the structure of Formula C, as describedherein.

In one embodiment, the present invention provides a method ofpotentiating the immune response to an infection in a subject with aninfection comprising contacting one or more cells of said subject with atherapeutically effective amount of a compound that promotes thedetachment of HK2 from mitochondria. In another embodiment, the presentinvention provides a method of potentiating the immune response to oneor more infectious diseases in a subject comprising contacting one ormore cells of said subject with a therapeutically effective amount of acompound that promotes the detachment of HK2 from mitochondria. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative. In one embodiment, thecompound is represented by the structure of Formula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof. In another embodiment,the compound is represented by the structure of Formula C, as describedherein.

In another embodiment, said infection is a microbial infection.

In one embodiment, the microbial infection is a bacterial infection.

In one embodiment, the bacterial infection comprises a Staphylococcusaureus infection.

In one embodiment, the bacterial infection comprises a Mycobacteriumtuberculosis infection.

In another embodiment, the N-acetylglucosamine (NAG) in thepeptidoglycan (PGN) of the bacterium is de-acetylated, therebydecreasing the inflammasome activation in response to infection by saidbacterium.

In one embodiment, the inflammasome activation is an NLRP3 inflammasomeactivation.

In another embodiment, said bacterial infection comprises a Bacillusanthracis infection.

In one embodiment, the microbial infection is a fungal infection.

In one embodiment, the microbial infection is a Plasmodium infection.

In one embodiment, the Plasmodium infection is a Plasmodium falciparum:infection.

In one embodiment, the Plasmodium infection comprises a Plasmodiumfalciparum infection. In another embodiment, the Plasmodium infectioncomprises a Plasmodium vivax, Plasmodium ovale, or Plasmodium malariaeinfection. In another embodiment, the Plasmodium infection comprises aPlasmodium knowlesi infection. In one embodiment, the Plasmodiuminfection causes malaria in said subject.

In another embodiment, said infection is a viral infection.

In one embodiment, the viral infection is a retroviral infection.

In one embodiment, the retroviral infection is a lentivirus infection.

In one embodiment, the lentivirus infection is a Human ImmunodeficiencyVirus (HIV) infection.

In another embodiment, said subject is afflicted with AfricanTrypanosomiasis (“sleeping sickness”), Cholera, Cryptosporidiosis,Dengue, Hepatitis A, Hepatitis B, Hepatitis C, Acquired ImmuneDeficiency Syndrome (AIDS), Influenza, Japanese Encephalitis,Leishmaniasis, Malaria, Measles, Meningitis, Onchocerciasis (“riverblindness”), Pneumonia, Rotavirus, Schistosomiasis, Shigellosis, StrepThroat, Tuberculosis, Typhoid, Yellow Fever, Ebola Hemorrhagic Fever orWest Nile Virus.

In another embodiment, said subject is immunocompromised.

In one embodiment, the subject has an immunodeficiency disorder.

In one embodiment, the immunodeficiency disorder is congenital(primary).

In one embodiment, the congenital immunodeficiency disorder comprisesX-linked agammaglobulinemia (XLA), common variable immunodeficiency(CVID), or severe combined immunodeficiency (SCID).

In one embodiment, the immunodeficiency disorder is acquired(secondary).

In one embodiment, the acquired immunodeficiency disorder comprisesimmunodeficiency resulting from a severe burn, chemotherapy, radiation,diabetes, malnutrition, AIDS, or an immune-complex disease.

In one embodiment, the immune-complex disease comprises viral hepatitis.

In one embodiment, the acquired immunodeficiency disorder comprises acancer of the immune system.

In one embodiment, the cancer of the immune system comprises leukemia ormultiple myeloma.

In one embodiment, the subject has a disease or condition linked to aprimary immunodeficiency disorder.

In one embodiment, the disease or condition comprisesataxia-telangiectasia, Chediak-Higashi syndrome, combinedimmunodeficiency disease, complement deficiencies, DiGeorge syndrome,hypogammaglobulinemia, Job syndrome, leukocyte adhesion defects,panhypogammaglobulinemia, Bruton's disease, congenitalagammaglobulinemia, selective deficiency of IgA, Wiskott-Aldrichsyndrome, HIV, or cancer therapy.

In one embodiment, the subject has a brain injury.

In another embodiment, contacting one or more cells of said subjectcomprises the step of administering a composition comprising saidcompound to said subject.

Pharmaceutical Compositions

Although the hexokinase 2 (HK2)/mitochondria-detaching compounds can beadministered alone, it is contemplated, in one embodiment, that thesecompounds will be administered in a pharmaceutical compositioncontaining the hexokinase 2 (HK2)/mitochondria-detaching compoundtogether with a pharmaceutically acceptable carrier or excipient.

In one embodiment, the active ingredient is a jasmonic derivative in thepharmaceutical composition is dissolved in any acceptable lipid carrier(e.g., fatty acids, oils to form, for example, a micelle or a liposome).In one embodiment, the composition additionally comprises at least oneother chemotherapeutic agent.

In one embodiment, at least one of the excipients is a non-aqueousexcipient.

In one embodiment, a compound as described herein which in oneembodiment is represented by the structure of Formula (VII), may bemixed with stabilizing excipients useful for topical administration in aformulation which is essentially free of compounds bearing anucleophilic character like water, alcoholic solvents and carboxylicacids. In some embodiments, the topical pharmaceutical composition ofthe present invention comprises a therapeutically effective amount of acompound as described herein, which in one embodiment is represented bythe structure of Formula (VII), or a pharmaceutically acceptable salt orsolvate thereof, and at least one pharmaceutically acceptable excipientselected from the group consisting of a glycol diester with a saturatedor unsaturated fatty acid, a triglyceride, an aprotic organic solvent,and any combination thereof, and optionally an oil and/or a wax.

In one embodiment, the pharmaceutically acceptable excipient comprises acompound represented by the structure of Formula (VIII):

-   -   wherein:    -   R¹, R² and R³ are each selected from the group consisting of H        and R⁴—COO—, wherein at least two of R¹, R² and R³ are R⁴—COO;        and    -   R⁴—COO— is the residue of a saturated or unsaturated fatty acid.

In another embodiment, the pharmaceutically acceptable excipientcomprises a triglyceride of Formula (VIII) or a combination thereofwherein R¹, R² and R³ are each R⁴—C(═O)—; and R⁴—C(═O)O— is the residueof an unsaturated or saturated fatty acid.

In some embodiments, the pharmaceutically acceptable excipient comprisesa triglyceride with a medium chain (e.g., C₈-C₁₂) saturated orunsaturated fatty acid. In one embodiment, the pharmaceuticallyacceptable excipient is caprylic/capric triglyceride (in one embodimentLabrafac lipophile WL 1349).

In another embodiment, the pharmaceutically acceptable excipientcomprises a propylene glycol diester with a saturated or unsaturatedfatty acid. In one embodiment, the fatty acid is a medium chain (e.g.,C₈-C₁₂) saturated or unsaturated fatty acid. In one embodiment, thepharmaceutically acceptable excipient is propylene glycol dicaprylate(in one embodiment, Labrafac PG).

In one embodiment, the organic solvent is dimethylsulfoxide (DMSO).

Compositions of the present invention may further comprise optionalcomponents such as lipid carriers such as oils or waxes so as to providea semi-solid consistency as desired. Additional inactive excipients suchas preservatives, anti-oxidants, perfumes and the like may also beadded.

Oils that can be used as optional components in the compositions of thepresent invention include, but are not limited to, petrolatum (petroleumjelly), vegetable oil, fruit oil, plant oil, animal oil, mineral oil,coconut oil, olive oil, lanolin, peanut oil, hydrogenated and sulphatedoils such as cottonseed oil, soybean oils, almond oil, sesame oil, andthe like. In another embodiment, the oil comprises corn oil, peanut oil,coconut oil, grape seed oil, sunflower oil, lemon oil, orange oil,peppermint oil, palm kernel oil, castor oil, hydrogenated cottonseedoil, hydrogenated soy oil, hydrogenated soybean oil, hydrogenatedvegetable oil, partially hydrogenated soybean oil, partiallyhydrogenated palm oil, hydrogenated castor oil, light mineral oil,mineral oil, or a combination thereof.

Waxes that can be used as optional components in the compositions of thepresent invention include, but are not limited to, paraffin wax,beeswax, carnauba wax, cetyl ester wax, microcrystalline wax, spermacetiwax, and the like.

In some embodiments, the composition of the invention comprises thecompound as described herein, propylene glycol dicaprylate (LabrafacPG), caprylic/capric triglyceride (Labrafac lipophile WL 1349),dimethylsulfoxide (DMSO), petrolatum and paraffin wax.

In some embodiments, the composition of the invention comprises acompound as described herein in an amount of from about 5% w/w to about30% w/w. In some embodiments, the composition of the invention comprisesa compound as described herein in an amount of about 5% w/w. In someembodiments, the composition of the invention comprises a compound asdescribed herein in an amount of about 10% w/w. In some embodiments, thecomposition of the invention comprises a compound as described herein inan amount of about 15% w/w. In some embodiments, the composition of theinvention comprises a compound as described herein in an amount of about20% w/w. In one embodiment, the compound is represented by the structureof Formula (VII).

In some embodiments, the composition of the invention comprises acompound as described herein in an amount of from about 5% to about 30%w/w; propylene glycol dicaprylate in an amount of from about 1% to about10% w/w; caprylic/capric triglyceride in an amount of from about 30% toabout 50% w/w; dimethyl sulfoxide in an amount of from about 0% to about10% w/w; petrolatum in an amount of from about 20% to about 50% w/w; andparaffin wax in an amount of from about 0 to about 10% w/w.

In some embodiments, the composition of the invention comprises acompound as described herein in an amount of about 5% w/w; propyleneglycol dicaprylate in an amount of about 10% w/w; caprylic/caprictriglyceride in an amount of about 50% w/w; dimethyl sulfoxide in anamount of about 1% w/w; petrolatum in an amount of about 30% w/w; andparaffin wax in an amount of about 4% w/w.

In other embodiments, the composition of the invention comprises acompound as described herein in an amount of about 10% w/w; propyleneglycol dicaprylate in an amount of about 7.5% w/w; caprylic/caprictriglyceride in an amount of about 47.5% w/w; dimethyl sulfoxide in anamount of about 1% w/w; petrolatum in an amount of about 30% w/w; andparaffin wax in an amount of about 4% w/w.

In other embodiments, the composition of the invention comprises: acompound as described herein in an amount of about 15% w/w; propyleneglycol dicaprylate in an amount of about 5% w/w; caprylic/caprictriglyceride in an amount of about 45% w/w; dimethyl sulfoxide in anamount of about 1% w/w; petrolatum in an amount of about 30% w/w; andparaffin wax in an amount of about 4% w/w.

In certain embodiments, the composition of the invention comprises acompound as described herein in an amount of about 20% w/w; propyleneglycol dicaprylate in an amount of about 2.5% w/w; caprylic/caprictriglyceride in an amount of about 42.5% w/w; dimethyl sulfoxide in anamount of about 1% w/w; petrolatum in an amount of about 30% w/w; andparaffin wax in an amount of about 4% w/w.

In some embodiments, the composition described above is essentially freeof water, alcoholic solvents and carboxylic acids.

The pharmaceutically acceptable excipients according to the principlesof the present invention may be a triglyceride, a glycol diester or acombination thereof. In some embodiments, the triglyceride and/or theglycol diester comprise saturated or unsaturated fatty acids. The term“saturated fatty acid” encompasses a carboxylic acid with an elongatedaliphatic saturated chain. The term “unsaturated fatty acid” encompassesa carboxylic acid with an elongated aliphatic chain which have one ormore double bonds between the carbon atoms. The length of the carbonchain may vary, but may generally be between 4 and 30 carbon atoms(C₄-C₃₀). In some embodiments, the triglyceride and/or glycol diestercomprises medium chain (e.g., C₈-C₁₂) saturated or unsaturated fattyacids.

The pharmaceutical compositions of the present invention can beformulated for administration by a variety of routes including oral,rectal, transdermal, parenteral (subcutaneous, intraperitoneal,intravenous, intraarterial, transdermal and intramuscular), topical,intranasal, or via a suppository. Such compositions are prepared in amanner well known in the pharmaceutical art and comprise as an activeingredient at least one compound of the present invention as describedhereinabove, and a pharmaceutically acceptable excipient or a carrier.

In one embodiment, the term “pharmaceutically acceptable” means approvedby a regulatory agency of the Federal or a state government or listed inthe U.S. Pharmacopeia or other generally recognized pharmacopeia for usein animals and, more particularly, in humans.

During the preparation of the pharmaceutical compositions according tothe present invention, the active ingredient is usually mixed with acarrier or excipient, which may be a solid, semi-solid, or liquidmaterial. The compositions can be in the form of tablets, pills,capsules, pellets, granules, powders, lozenges, sachets, cachets,elixirs, suspensions, dispersions, emulsions, solutions, syrups,aerosols (as a solid or in a liquid medium), ointments containing, forexample, up to 10% by weight of the active compound, soft and hardgelatin capsules, suppositories, sterile injectable solutions, andsterile packaged powders.

The carriers may be any of those conventionally used and are limitedonly by chemical-physical considerations, such as solubility and lack ofreactivity with the compound for use in the present invention, and bythe route of administration. The choice of carrier will be determined bythe particular method used to administer the pharmaceutical composition.Some examples of suitable carriers include lactose, glucose, dextrose,sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate,alginates, tragacanth, gelatin, calcium silicate, microcrystallinecellulose, polyvinylpyrrolidone, cellulose, water and methylcellulose.The formulations can additionally include lubricating agents such astalc, magnesium stearate, and mineral oil; wetting agents, surfactants,emulsifying and suspending agents; preserving agents such as methyl- andpropylhydroxybenzoates; sweetening agents; flavoring agents, colorants,buffering agents (e.g., acetates, citrates or phosphates),disintegrating agents, moistening agents, antibacterial agents,antioxidants (e.g., ascorbic acid or sodium bisulfite), chelating agents(e.g., ethylenediaminetetraacetic acid), and agents for the adjustmentof tonicity such as sodium chloride. Other pharmaceutical carriers canbe sterile liquids, such as water and oils, including those ofpetroleum, animal, vegetable or synthetic origin, such as peanut oil,soybean oil, mineral oil, sesame oil and the like, polyethylene glycols,glycerine, propylene glycol or other synthetic solvents. In oneembodiment, water is a preferred carrier when the pharmaceuticalcomposition is administered intravenously. Saline solutions and aqueousdextrose and glycerol solutions can also be employed as liquid carriers,particularly for injectable solutions.

For preparing solid compositions such as tablets, the principal activeingredient is mixed with a pharmaceutical excipient to form a solidpreformulation composition containing a homogeneous mixture of acompound of the present invention. When referring to thesepreformulation compositions as homogeneous, it is meant that the activeingredient is dispersed evenly throughout the composition so that thecomposition may be readily subdivided into equally effective unit dosageforms such as tablets, pills and capsules. This solid preformulation isthen subdivided into unit dosage forms of the type described abovecontaining from, for example, 0.1 to about 500 mg of the activeingredient of the present invention.

Any method can be used to prepare the pharmaceutical compositions. Soliddosage forms can be prepared by wet granulation, dry granulation, directcompression and the like.

The solid dosage forms of the present invention may be coated orotherwise compounded to provide a dosage form affording the advantage ofprolonged action. For example, the tablet or pill can comprise an innerdosage and an outer dosage component, the latter being in the form of anenvelope over the former. The two components can be separated by anenteric layer, which serves to resist disintegration in the stomach andpermit the inner component to pass intact into the duodenum or to bedelayed in release. A variety of materials can be used for such entericlayers or coatings, such materials including a number of polymeric acidsand mixtures of polymeric acids with such materials as shellac, cetylalcohol, and cellulose acetate.

The liquid forms in which the compositions of the present invention maybe incorporated, for administration orally or by injection, includeaqueous solutions, suitably flavored syrups, aqueous or oil suspensions,and flavored emulsions with edible oils such as cottonseed oil, sesameoil, coconut oil, or peanut oil, as well as elixirs and similarpharmaceutical vehicles.

Compositions for inhalation or insulation include solutions andsuspensions in pharmaceutically acceptable aqueous or organic solvents,or mixtures thereof, and powders. The liquid or solid compositions maycontain suitable pharmaceutically acceptable excipients as describedabove. In one embodiment, the compositions are administered by the oralor nasal respiratory route for local or systemic effect. Compositions inpharmaceutically acceptable solvents may be nebulized by use of inertgases. Nebulized solutions may be breathed directly from the nebulizingdevice or the nebulizing device may be attached to a face masks tent, orintermittent positive pressure breathing machine. Solution, suspension,or powder compositions may be administered, orally or nasally, fromdevices that deliver the formulation in an appropriate manner.

Another formulation employed in the methods of the present inventionemploys transdermal delivery devices (“patches”). Such transdermalpatches may be used to provide continuous or discontinuous infusion ofthe compounds of the present invention in controlled amounts. Theconstruction and use of transdermal patches for the delivery ofpharmaceutical agents is well known in the art.

In yet another embodiment, the composition is prepared for topicaladministration. In one embodiment, the topical composition comprises anointment. In another embodiment, the topical composition comprises acream. In another embodiment, the topical composition comprises alotion. In another embodiment, the topical composition comprises a gel.In another embodiment, the topical composition comprises a paste. Inanother embodiment, the topical composition comprises a drop. In anotherembodiment, the topical composition comprises a foam. For topicaladministration to body surfaces using, for example, creams, gels, drops,ointments and the like, the compounds of the present invention can beprepared and applied in a physiologically acceptable diluent with orwithout a pharmaceutical carrier.

In one embodiment, the present invention may be used topically ortransdermally to treat cancer, for example, melanoma. Adjuvants fortopical or gel base forms may include, for example, sodiumcarboxymethylcellulose, polyacrylates,polyoxyethylene-polyoxypropylene-block polymers, polyethylene glycol andwood wax alcohols.

Alternative formulations include nasal sprays, liposomal formulations,slow-release formulations, controlled-release formulations and the like,as are known in the art.

In one embodiment, the compositions are formulated in a unit dosageform. The term “unit dosage forms” refers to physically discrete unitssuitable as unitary dosages for human subjects and other mammals, eachunit containing a predetermined quantity of active material calculatedto produce the desired therapeutic effect, in association with asuitable pharmaceutical excipient.

In preparing a formulation, it may be necessary to mill the activeingredient to provide the appropriate particle size prior to combiningwith the other ingredients. If the active compound is substantiallyinsoluble, it ordinarily is milled to a particle size of less than 200mesh. If the active ingredient is substantially water soluble, theparticle size is normally adjusted by milling to provide a substantiallyuniform distribution in the formulation, e.g. about 40 mesh.

It may be desirable to administer the pharmaceutical composition of theinvention locally to the area in need of treatment; this may be achievedby, for example, and not by way of limitation, local infusion duringsurgery, topical application, e.g., in conjunction with a wound dressingafter surgery, by injection, by means of a catheter, by means of asuppository, or by means of an implant, said implant being of a porous,non-porous, or gelatinous material. According to some embodiments,administration can be by direct injection e.g., via a syringe, at thesite of a tumor or neoplastic or pre-neoplastic tissue.

The compounds may also be administered by any convenient route, forexample by infusion or bolus injection, by absorption through epitheliallinings (e.g., oral mucosa, rectal and intestinal mucosa, etc.), and maybe administered together with other therapeutically active agents. Inone embodiment, administration is localized. In another embodiment,administration is systemic. In addition, it may be desirable tointroduce the pharmaceutical compositions of the invention into thecentral nervous system by any suitable route, including intraventricularand intrathecal injection; intraventricular injection may be facilitatedby an intraventricular catheter, for example, attached to a reservoir.Pulmonary administration can also be employed, e.g., by use of aninhaler or nebulizer, and formulation with an aerosolizing agent.

A compound of the present invention can be delivered in an immediaterelease or in a controlled release system. In one embodiment, aninfusion pump may be used to administer a compound of the invention,such as one that is used for delivering chemotherapy to specific organsor tumors (see Buchwald et al., 1980, Surgery 88: 507; Saudek et al.,1989, N. Engl. J. Med. 321: 574). In one embodiment, a compound of theinvention is administered in combination with a biodegradable,biocompatible polymeric implant, which releases the compound over acontrolled period of time at a selected site. Examples of polymericmaterials include polyanhydrides, polyorthoesters, polyglycolic acid,polylactic acid, polyethylene vinyl acetate, copolymers and blendsthereof (See, Medical applications of controlled release, Langer andWise (eds.), 1974, CRC Pres., Boca Raton, Fla.). In yet anotherembodiment, a controlled release system can be placed in proximity ofthe therapeutic target, thus requiring only a fraction of the systemicdose.

Furthermore, at times, the pharmaceutical compositions may be formulatedfor parenteral administration (subcutaneous, intravenous, intraarterial,transdermal, intraperitoneal or intramuscular injection) and may includeaqueous and non-aqueous, isotonic sterile injection solutions, which cancontain anti-oxidants, buffers, bacteriostats, and solutes that renderthe formulation isotonic with the blood of the intended recipient, andaqueous and non-aqueous sterile suspensions that include suspendingagents, solubilizers, thickening agents, stabilizers, and preservatives.Oils such as petroleum, animal, vegetable, or synthetic oils and soapssuch as fatty alkali metal, ammonium, and triethanolamine salts, andsuitable detergents may also be used for parenteral administration. Theabove formulations may also be used for direct intra-tumoral injection.Further, in order to minimize or eliminate irritation at the site ofinjection, the compositions may contain one or more nonionicsurfactants. Suitable surfactants include polyethylene sorbitan fattyacid esters, such as sorbitan monooleate and the high molecular weightadducts of ethylene oxide with a hydrophobic base, formed by thecondensation of propylene oxide with propylene glycol.

The parenteral formulations can be presented in unit-dose or multi-dosesealed containers, such as ampoules and vials, and can be stored in afreeze-dried (lyophilized) condition requiring only the addition of thesterile liquid carrier, for example, water, for injections, immediatelyprior to use. Extemporaneous injection solutions and suspensions can beprepared from sterile powders, granules, and tablets of the kindpreviously described and known in the art.

Alternatively, the jasmonate derivatives of the present invention can beused in hemodialysis such as leukophoresis and other related methods,e.g., blood is drawn from the patient by a variety of methods suchdialysis through a column/hollow fiber membrane, cartridge etc, istreated with the jasmonate derivatives Ex-vivo, and returned to thepatient following treatment. Such treatment methods are well known anddescribed in the art. See, e.g., Kolho et al. (J. Med. Virol. 1993,40(4): 318-21); Ting et al. (Transplantation, 1978, 25(1): 31-3); thecontents of which are hereby incorporated by reference in theirentirety.

As used herein, the term “administering” refers to bringing in contactwith a compound of the present invention. In one embodiment, thecompositions are applied locally. In another embodiment, thecompositions are applied systemically. Administration can beaccomplished to cells or tissue cultures, or to living organisms, forexample humans.

A “therapeutic” treatment is a treatment administered to a subject whoexhibits signs of pathology for the purpose of diminishing oreliminating those signs.

In one embodiment, compounds and compositions of the present inventionsare provided to a subject in a therapeutically effective amount. In oneembodiment, a “therapeutically effective amount” of a compound of theinvention is that amount of compound which is sufficient to provide abeneficial effect to the subject to which the compound is administered.

In one embodiment, “treating” refers to both therapeutic treatment andprophylactic or preventative measures, wherein the object is to preventor lessen the targeted pathologic condition or disorder as describedhereinabove. Thus, in one embodiment, treating may include directlyaffecting or curing, suppressing, inhibiting, preventing, reducing theseverity of, delaying the onset of, reducing symptoms associated withthe disease, disorder or condition, or a combination thereof. Thus, inone embodiment, “treating” refers inter alia to delaying progression,expediting remission, inducing remission, augmenting remission, speedingrecovery, increasing efficacy of or decreasing resistance to alternativetherapeutics, or a combination thereof. In one embodiment, “preventing”refers, inter alia, to delaying the onset of symptoms, preventingrelapse to a disease, decreasing the number or frequency of relapseepisodes, increasing latency between symptomatic episodes, or acombination thereof. In one embodiment, “suppressing” or “inhibiting”,refers inter alia to reducing the severity of symptoms, reducing theseverity of an acute episode, reducing the number of symptoms, reducingthe incidence of disease-related symptoms, reducing the latency ofsymptoms, ameliorating symptoms, reducing secondary symptoms, reducingsecondary infections, prolonging patient survival, or a combinationthereof.

According to any of the methods of the present invention and in oneembodiment, the subject is human. In another embodiment, the subject ismammalian. In another embodiment, the subject is a primate, which in oneembodiment, is a non-human primate. In another embodiment, the subjectis murine, which in one embodiment is a mouse, and, in anotherembodiment is a rat. In another embodiment, the subject is canine,feline, bovine, equine, caprine, ovine, porcine, simian, ursine,vulpine, or lupine. In one embodiment, the subject is a chicken or fish.

Doses and Routes of Administration

In one embodiment, the composition is formulated for parenteraladministration.

In another embodiment, the composition is formulated for intravenous,subcutaneous, or intramuscular administration. In another embodiment,the composition is formulation for per os (P.O.) or oral administration.In one embodiment, is formulated for topical administration.

In one embodiment, the jasmonic derivatives as described herein areformulated for subcutaneous injection.

In one embodiment, the piperazine derivatives, as described herein areformulated for oral administration.

In one embodiment, the composition comprises 5% of the compound asdescribed herein. In another embodiment, the composition comprises 10%of the compound as described herein. In another embodiment, thecomposition comprises 15% of the compound as described herein. Inanother embodiment, the composition comprises 20% of the compound asdescribed herein. In another embodiment, the composition comprises 30%of the compound as described herein. In another embodiment, thecomposition comprises 40% of the compound as described herein. Inanother embodiment, the composition comprises 50% of the compound asdescribed herein. In one embodiment, the compound is the compoundrepresented by the structure of Formula (VII).

In a further embodiment, the compound as described herein is formulatedas 250 mg of a 5%, 10%, 20%, 30%, 40% or 50% cream, gel, ointment orpaste. In one embodiment, the compound is the compound represented bythe structure of Formula (VII).

In another embodiment, the compound as described herein is formulated as200 mg of a 5%, 10%, 20%, 30%, 40% or 50% cream, gel, ointment or paste.In one embodiment, the compound is the compound represented by thestructure of Formula (VII).

In one embodiment, the compound as described herein is administered onceper day. In another embodiment, the compound as described herein isadministered twice per day. In another embodiment, the compound asdescribed herein is administered three times per day. In anotherembodiment, the compound as described herein is administered four timesper day. In another embodiment, the compound as described herein isadministered once every two days, once every three days, twice a week,once a week, once every 2 weeks, once every 3 weeks. In one embodiment,the compound is the compound represented by the structure of Formula(VII). In another embodiment the compound is the compound represented bythe structure of Formula (II), which in one embodiment comprises thecompounds of Formula (20-24).

In one embodiment, the compound as described herein is administered for7 days to 28 days. In another embodiment, the compound as describedherein is administered for 7 days to 8 weeks. In another embodiment, thecompound as described herein is administered for 7 days to 50 days. Inanother embodiment, the compound as described herein is administered for7 days to six months. In another embodiment, the compound as describedherein is administered for 7 days to one and half years. In anotherembodiment, the compound as described herein is administered for 14 daysto 12 months. In another embodiment, the compound as described herein isadministered for 14 days to 3 years. In another embodiment, the compoundas described herein is administered for several years. In anotherembodiment, the compound as described herein is administered for onemonth to six months. In one embodiment, the compound is the compoundrepresented by the structure of Formula (VII). In another embodiment thecompound is the compound represented by the structure of Formula (II),which in one embodiment comprises the compounds of Formula (20-24).

In one embodiment, the compound as described herein is administered for7 days. In another embodiment, the compound as described herein isadministered for 14 days. In another embodiment, the compound asdescribed herein is administered for 21 days. In another embodiment, thecompound as described herein is administered for 28 days. In anotherembodiment, the compound as described herein is administered for 50days. In another embodiment, the compound as described herein isadministered for 56 days. In another embodiment, the compound asdescribed herein is administered for 84 days. In another embodiment, thecompound as described herein is administered for 90 days. In anotherembodiment, the compound as described herein is administered for 120days. In one embodiment, the compound is the compound represented by thestructure of Formula (VII). In another embodiment the compound is thecompound represented by the structure of Formula (II), which in oneembodiment comprises the compounds of Formula (20-24).

The amount of a compound of the invention that will be effective in thetreatment of a particular disorder or condition, including cancer, willdepend on the nature of the disorder or condition, and can be determinedby standard clinical techniques. In addition, in vitro assays mayoptionally be employed to help identify optimal dosage ranges. Theprecise dose to be employed in the formulation will also depend on theroute of administration, and the seriousness of the disease or disorder,and should be decided according to the judgment of the practitioner andeach patient's circumstances. In one embodiment, the dosage will bewithin the range of 0.01-1000 mg/kg of body weight, in anotherembodiment, 0.1 mg/kg to 100 mg/kg, in another embodiment, 1 mg/kg to 10mg/kg, and, in another embodiment, 100-400 mg/kg. Effective doses may beextrapolated from dose-response curves derived from in vitro or animalmodel test bioassays or systems.

As described hereinabove, in one embodiment, methods of the presentinvention further comprise the step of contacting one or more cells ofsaid subject with a therapeutic agent. In one embodiment, thechemotherapeutic agent is administered prior to administration thecompound as described herein. In another embodiment, thechemotherapeutic agent is administered concurrently with the compoundrepresented by the structure as described herein. In another embodiment,the chemotherapeutic agent is administered after administration thecompound as described herein. In one embodiment, the compound is thecompound represented by the structure of Formula (VII). In anotherembodiment the compound is the compound represented by the structure ofFormula (II), which in one embodiment comprises the compounds of Formula(20-24).

In another embodiment, the present invention provides a method ofinducing cell pyroptosis in a subject comprising contacting one or morecells of said subject with a therapeutically effective amount of acompound that promotes the detachment of HK2 from mitochondria. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative, which in one embodiment, isrepresented by the structure of Formula (II). In one embodiment, thecompound is represented by the structure of Formula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof. In another embodiment,the compound is represented by the structure of Formula C, as describedherein.

In one embodiment, pyroptosis is a form of cell death that is distinctfrom apoptosis.

In one embodiment, administration of an HK2/mitochondria-detachingcompound as described herein leads to dissociation of HK2 from VDAC. Inanother embodiment, administration of an HK2/mitochondria-detachingcompound leads to activation of an inflammasome, which, in oneembodiment, leads to an immune response directed to cancer cells. Inanother embodiment, administration of the compound of anHK2/mitochondria-detaching compound leads to apoptosis, which in oneembodiment, leads to the death of cancer cells. In another embodiment,administration of an HK2/mitochondria-detaching compound leads toreduced glycolysis. In one embodiment, the HK2/mitochondria-detachingcompound comprises a jasmonic acid derivative. In one embodiment, thejasmonic acid derivative comprises a compound represented by thestructure of Formula (VII) as described herein. In another embodiment,the jasmonic acid derivative comprises a compound represented by thestructure of Formula C, as described herein. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative.

In one embodiment, stereoisomers of the jasmonate ester derivatives arecontemplated either in admixture or in pure or substantially pure form.The jasmonate derivatives can have asymmetric centers at any of theatoms. Consequently, the compounds can exist in enantiomeric ordiastereomeric forms or in mixtures thereof. The present inventioncontemplates the use of any racemates (i.e. mixtures containing equalamounts of each enantiomers), enantiomerically enriched mixtures (i.e.,mixtures enriched for one enantiomer), pure enantiomers ordiastereomers, or any mixtures thereof. The chiral centers can bedesignated as R or S or R, S, D or d or L or 1 or d,l or D,L. Inaddition, several of the compounds of the invention contain one or moredouble bonds. The present invention intends to encompass all structuraland geometrical isomers including cis, trans, E and Z isomers,independently at each occurrence.

One or more of the compounds of the invention, may be present as a salt.The term “salt” encompasses both basic and acid addition salts,including but not limited to, carboxylate salts or salts with aminenitrogens, and include salts formed with the organic and inorganicanions and cations discussed below. Furthermore, the term includes saltsthat form by standard acid-base reactions with basic groups (such asamino groups) and organic or inorganic acids. Such acids includehydrochloric, hydrofluoric, trifluoroacetic, sulfuric, phosphoric,acetic, succinic, citric, lactic, maleic, fumaric, palmitic, cholic,pamoic, mucic, D-glutamic, D-camphoric, glutaric, phthalic, tartaric,lauric, stearic, salicylic, methanesulfonic, benzenesulfonic, sorbic,picric, benzoic, cinnamic, and like acids.

The term “organic or inorganic cation” refers to counter-ions for thecarboxylate anion of a carboxylate salt. The counter-ions are chosenfrom the alkali and alkaline earth metals (such as lithium, sodium,potassium, barium, aluminum and calcium); ammonium and mono-, di- andtri-alkyl amines such as trimethylamine, cyclohexylamine; and theorganic cations, such as dibenzylammonium, benzylammonium,2-hydroxyethylammonium, bis(2-hydroxyethyl)ammonium,phenylethylbenzylammonium, dibenzylethylenediammonium, and like cations.See, for example, Berge et al., J. Pharm. Sci., 66: 1-19, 1977. Othercations encompassed by the above term include the protonated form ofprocaine, quinine and N-methylglucosamine, and the protonated forms ofbasic amino acids such as glycine, ornithine, histidine, phenylglycine,lysine and arginine. Furthermore, any zwitterionic form of the instantcompounds formed by a carboxylic acid and an amino group is alsocontemplated.

The present invention also includes solvates of the compounds of thepresent invention and salts thereof. “Solvate” means a physicalassociation of a compound of the invention with one or more solventmolecules. This physical association involves varying degrees of ionicand covalent bonding, including hydrogen bonding. In certain instances,the solvate will be capable of isolation. “Solvate” encompasses bothsolution-phase and isolatable solvates. Non-limiting examples ofsuitable solvates include ethanolates, methanolates and the like.“Hydrate” is a solvate wherein the solvent molecule is water.

The present invention also includes polymorphs of the compounds of thepresent invention and salts thereof. The term “polymorph” refers to aparticular crystalline state of a substance, which can be characterizedby particular physical properties such as X-ray diffraction, IR or Ramanspectra, melting point, and the like.

Kits

In another embodiment, the present invention provides a kit foractivating an immune response comprising a therapeutically effectiveamount of a compound that promotes the detachment of HK2 frommitochondria. In one embodiment, the HK2/mitochondria-detaching compoundcomprises a jasmonic acid derivative. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative,which in one embodiment, is represented by the structure of Formula(II). In one embodiment, the compound is represented by the structure ofFormula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof; and instructions foruse for activating an inflammatory response, as described herein. Inanother embodiment, the compound is represented by the structure ofFormula C, as described herein.

In another embodiment, the present invention provides a kit foractivating a cell-mediated immune response comprising a therapeuticallyeffective amount of a compound that promotes the detachment of HK2 frommitochondria. In one embodiment, the HK2/mitochondria-detaching compoundcomprises a jasmonic acid derivative. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative,which in one embodiment, is represented by the structure of Formula(II). In one embodiment, the compound is represented by the structure ofFormula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof; and instructions foruse for activating an inflammatory response, as described herein. Inanother embodiment, the compound is represented by the structure ofFormula C, as described herein.

In another embodiment, the present invention provides a kit foractivating an inflammatory response comprising a therapeuticallyeffective amount of a compound that promotes the detachment of HK2 frommitochondria. In one embodiment, the HK2/mitochondria-detaching compoundcomprises a jasmonic acid derivative. In another embodiment, theHK2/mitochondria-detaching compound comprises a piperazine derivative,which in one embodiment, is represented by the structure of Formula(II). In one embodiment, the compound is represented by the structure ofFormula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof; and instructions foruse for activating an inflammatory response, as described herein. Inanother embodiment, the compound is represented by the structure ofFormula C, as described herein.

In another embodiment, the present invention provides a kit foractivating an inflammasome-mediated immune response comprising atherapeutically effective amount of a compound that promotes thedetachment of HK2 from mitochondria. In one embodiment, theHK2/mitochondria-detaching compound comprises a jasmonic acidderivative. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative, which in one embodiment, isrepresented by the structure of Formula (II). In one embodiment, thecompound is represented by the structure of Formula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof; and instructions foruse for activating an inflammasome-mediated immune response, asdescribed herein. In another embodiment, the compound is represented bythe structure of Formula C, as described herein.

In another embodiment, the present invention provides a kit for treatinga cancer comprising therapeutically effective amount of a compound thatpromotes the detachment of HK2 from mitochondria. In one embodiment, theHK2/mitochondria-detaching compound comprises a jasmonic acidderivative. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative, which in one embodiment, isrepresented by the structure of Formula (II). In one embodiment, thecompound is represented by the structure of Formula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₁₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₂ alkyl, unsubstituted or substitutedC₁-C₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof, and instructions foruse for treating said cancer, as described herein. In anotherembodiment, the kit comprises additional containers comprisingadditional cancer therapies as described herein. In another embodiment,the compound is represented by the structure of Formula C, as describedherein.

In another embodiment, the present invention provides a kit for treatingan infection comprising therapeutically effective amount of a compoundthat promotes the detachment of HK2 from mitochondria. In oneembodiment, the HK2/mitochondria-detaching compound comprises a jasmonicacid derivative. In another embodiment, the HK2/mitochondria-detachingcompound comprises a piperazine derivative, which in one embodiment, isrepresented by the structure of Formula (II). In one embodiment, thecompound is represented by the structure of Formula (VII):

wherein A is COR¹; R¹ is an unsubstituted or substituted heteroaryloxy;R² is selected from the group consisting of hydrogen, unsubstituted orsubstituted C₁-C₂ alkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸, oxo and NR^(9a)R^(9b); R³, R⁴, R⁵, R⁶ and R⁷ are eachindependently selected from the group consisting of hydrogen, halogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₁-C₁₂ haloalkyl, unsubstituted or substituted C₃-C₈ cycloalkyl,unsubstituted or substituted aryl, unsubstituted or substitutedheteroaryl, OR⁸ and NR^(9a)R^(9b), or R⁵ and R⁶ together with thecarbons to which they are attached form a C₃-C₈ cycloalkyl or a C₃-C₈cycloalkyl substituted by halo; or one of R⁵ and R⁶ represents an oxygenatom which is bonded to C₆, thereby forming an oxygen-containing 6 or 5membered heterocyclic ring, respectively; wherein the bond between C₉and C₁₀ can be a single or double bond; R⁸, R^(9a) and R^(9b) are eachindependently selected from the group consisting of hydrogen,unsubstituted or substituted C₁-C₁₂ alkyl, unsubstituted or substitutedC₃-C₈ cycloalkyl, unsubstituted or substituted aryl, unsubstituted orsubstituted heteroaryl, glucosyl, or R^(9a) and R^(9b) can together withthe nitrogen to which they are attached form an unsubstituted orsubstituted heterocyclic or heteroaromatic ring optionally containingone or more additional heteroatom selected from O, N and S; R¹⁵ and R¹⁶are each hydrogen; and n is selected from 0, 1 and 2; including salts,hydrates, solvates, polymorphs, optical isomers, geometrical isomers,enantiomers, diastereomers, and mixtures thereof; and instructions foruse to treat an infection as described herein. In another embodiment,the compound is represented by the structure of Formula C, as describedherein.

In one embodiment, the kit as described herein comprises a jasmonicderivative formulated for subcutaneous administration. In oneembodiment, the kit comprises two vials, wherein the first vial containsthe jasmonic derivative and the second vial contains a solvent suitablefor subcutaneous injections, which in one embodiment comprises ahydrophobic solvent and, in another embodiment, comprises a mixedhydrophobic/polar solvent. In one embodiment, the content of the vialswill be mixed by the nurse shortly before administration.

In one embodiment, the kit comprises additional containers comprisingadditional cancer therapeutics as described herein.

Any patent, patent application publication, or scientific publication,cited herein, is incorporated by reference herein in its entirety.

The following examples are presented in order to more fully illustratethe uses described herein.

EXAMPLES Example 1 Topical Formula C Treatment in the Mouse B16-F10Melanoma Model Inhibits Tumor Growth and Increases Inflammation

Cell Lines

Mouse melanoma (B16-F10) cells were used. Mouse B16-F10 melanoma cellswere maintained in DMEM. Media was supplemented with 10%heat-inactivated FCS, penicillin (100 U/ml), streptomycin (100 μg/ml),Sodium pyruvate (2 mM), and L-glutamine (4 mM). Confluent cultures werewashed with PBS, detached with trypsin (0.25%), centrifuged, andsubcultured in 96-well microtiter test plates.

Animal Studies

Female C57B1/6 mice at 8 weeks of age were purchased from Harlan,Jerusalem, Israel. The animals were housed (6 or 7/cage) undercontrolled conditions (temperature, light, humidity) and given food andwater ad libitum. After 14 days of acclimatization, the animals wereinoculated with 0.25*10⁶ B16-F10 melanoma cells/mouse in 30 μl PBS underthe skin of the right ear of each mouse.

The mice were inoculated with B16-F10 melanoma cells and dividedrandomly into three groups (9 mice each): a) B16-F10 inoculated,untreated; and b) B16-F10 inoculated, Formula C (50% v/v in olive oil)treated

The treatment was initiated when the tumors reached the target volume ofapproximately 1 mm³. Drug was topically administered to the area of thetumor daily for 18 days.

Inflammation Score

Inflammation was graded on a scale of 0-4 as: 0 (absence ofinflammation), 1 (redness), 2 (tissue inflammation), 3 (pronouncedtissue inflammation), and 4 (damage to the ear).

Results

Untreated, B16-F10-incoluated mice demonstrated a steadily increasingtumor volume over the first 18 days of treatment (FIG. 1). In contrast,mice treated with 50% Formula C showed no tumor growth for 11 days, andthereafter only slight increase in tumor volume, which was significantlylower than the tumor volume in the untreated group (FIG. 1).

There is evidence to suggest the inflammatory system may inhibit thedevelopment of cancer, possibly via cancer-associated recognitionevents. The host may have a dedicated mechanism to perceive andeliminate transformed cells. In addition, adaptive immune recognition oftumor-associated and specific antigens also may be an important means bywhich the immune and inflammatory systems control the development ofcancer.

Therefore, the effect of Formula C treatment on ear inflammation inB16-F10 melanoma mice was examined. Untreated B16-F10 melanoma mice didnot exhibit any significant inflammation, as determined by inflammationscore (inflammation score of 0; FIG. 2). In contrast, B16-F10 melanomamice treated topically with Formula C exhibited increased inflammationover the course of treatment, attaining an average inflammation score ofabout 3 by about the 15^(th) day of treatment and increasing to over 3before the 18^(th) day of treatment (FIG. 2).

Formula C is therefore effective in limiting tumor growth and increasesinflammation, which may promote an effective immune response againsttumor cells.

Example 2 Topical Formula C Treatment in the AK/cSCC Mouse ModelTriggers Immune Response

Topical Formulations

The formulation ingredients for vehicle, 2.5%, and 5% Formula C ointmentconsisted of: 32% Gelucire44/14® (Gattefossé, Saint-Priest, France), and68% combined Labrasol® (Gattefossé, Saint-Priest, France) withcorresponding amount of Formula C (w/w). New ointments were preparedevery 2 weeks. The 40% topical solution was prepared just prior to useby mixing (w/w) 60% propylene glycol (Sigma, St. Louis, Mo., USA) with40% Formula C.

Animal Studies

Female SKH-1 mice were obtained from Charles River Laboratories(Wilmington, N.C.). Mice were treated and managed at Pharmaseed Ltd.(Ness Ziona, Israel).

Experimental Procedure

UVB-induced skin damage was adopted from previous publications.Forty-eight SKH-1 hairless female mice were chronically exposed to 1.25times the Minimal Erythemal Dose (MED) of UVB radiation. Lamp VL-6.M wasused, irradiating at 312 nm with an intensity of 0.89 mW/cm2 (validatedon Day 44) at a distance of 15 cm. Determination of UVB irradiationMinimal Erythemal Dose (MED) was performed prior to the beginning of thestudy. Animals were UVB irradiated for different duration (starting at30 sec) and the presence of erythema was recorded 24 and 48 hours later.1.25 times the MED of UVB irradiation was established to be equal to anexposure time of 5 min. Mice were UVB irradiated for 4-5 min at eachirradiation session during the 16 weeks of pre-treatment diseaseinduction. Prior to every irradiation session, the lamp was turned onfor 5 min to ensure it reaches its maximal intensity. The irradiationschedule was five times a week for two weeks, one week off, and thenthree times a week through week 16. By that time, more than 60% of theanimals had developed at least one SCC lesion. Three days after thefinal irradiation dose, the mice were randomly assigned to 1 of 4treatment groups (N=12 mice/group), with similar tumor burdendistribution between groups, and a 50-day treatment phase started. The 6cm² (2×3 cm) treatment area was marked on the back of the mice with ablack eight points rectangle tattoo. Five spare mice with the highesttumor burden were excluded from the main study. Treatment groupsincluded: vehicle, 2.5% or 5% Formula C ointment, 50 μl of which wasapplied once-daily for 50 days, or 50 μl of 40% Formula C topicalsolution applied once-daily for the first 5 days of every three-weekcycle (total of three treatment cycles). Following administration, eachanimal was placed in individual cage without bedding for 1.5 hours,until the ointment was absorbed.

IHC, Histology and Semi-Quantitative Analysis of IHC Slides

Histopathological analysis and IHC was performed according to standardprotocols. Primary antibodies used: Rabbit anti-CD3 (SP7), rabbitanti-F4/80 (SB-M3072, SB-M4152, Nordic Biosite, Tiiby, Sweden), mouseanti-Langerin/CD207 (DDX0361, Dendritics, Lyon, France). A polymersystem was used as a secondary antibody (KDB-10046 and KDB-10007, NordicBiosite, Tiiby, Sweden). Semi-quantitative analysis was done in ablinded manner by a single experienced pathologist scoring ×40 highpower fields, 7 different fields per slide.

Statistical Analysis

Statistical analysis was carried out using Graphpad Prism version 5.03(Graphpad Software, San Diego, Calif.).

Results

Histopathology H&E staining established the presence of mildinflammation in all skin samples that were exposed to prolonged UVBradiation (compared to skin of naïve control mice), without apparentdifference in overall inflammation between vehicle-treated and FormulaC-treated groups. However, a detailed analysis of the distribution ofvarious immune cells in the treated skin revealed statisticallysignificant changes. A statistically significant, dose-related increasein mast cells was observed in the skin of Formula C-treated micecompared to vehicle-treated mice (FIG. 3A). A statistically-significantincrease was also observed for CD3+ T cells (FIG. 3B) and for Langerin+dendritic cells (FIG. 3C) between the vehicle-treated group and theFormula C high dose (40%) group, trending also at lower doses. There wasno change in F4/80 general staining for macrophages (data not shown).

Example 3 Formula C Treatment Induced IL-1β Secretion from ActivatedMacrophages

The human monocyte cell line, THP-1, was grown in 10% FCS in RPMImedium. Cells were plated in 24 well plate, 5*10⁶ cells/well and weredifferentiated to macrophages in the presence of 0.5 uM PMA (phorbol12-myristate 13-acetate) for 24 hr. Media was replaced to contain LPS(10 ug/mL), 24 hr stimulation. ATP was added for 2 hours. Supernatantswere collected and assayed by ELISA for the presence of IL-1β.

Formula C treatment induced IL-1β secretion from human activatedmacrophages (FIG. 4). This strongly supports the hypothesis that FormulaC activates the NLRP3 inflammasome which results in IL-1β secretion andstimulation of the immune system.

BALB/C mouse bone marrow-derived macrophages (BMDM) or C57BL mouse bonemarrow-derived macrophages were seeded at 3×10⁴ cells/well in 96 wellplates. The following day, cells were stimulated with interferon-gamma(IFNγ) overnight 50 ng/ml. Then Ultra-pure LPS (100 ng/ml) was added for4 hours, followed by compound treatment for overnight, except for ATP(all compounds were added on top); ATP for 40 min. ELISA was performedon supernatants for mouse IL-1β detection. ATP and 2-DG were used aspositive controls. No signal without IFN-γ even when seeded at 100,000cells per well.

Formula C dose-dependently elicited IL-1β secretion from primaryactivated ex vivo macrophages (FIGS. 5A, 5C). Formula C had littleeffect on cell viability, even at high dosage (FIGS. 5B, 5D). Thisstrongly supports the hypothesis that Formula C activates the NLRP3inflammasome which results in IL-1J secretion and stimulation of theimmune system.

Colorectal cancer was induced in Balb/C mice using CT26 mouse colorectalcancer cells. Formula C was administered SC with esterase-inhibitor BNPP(IP).

Administration of Formula C decreased tumor volume in Balb/C mice in asyngeneic cancer model (CT26 mouse colorectal cancer (FIG. 6A)). FormulaC administration shifts macrophages in vivo from pro-tumor (M2)macrophages to antitumor (M1) macrophages.

Pro-tumor M2 Macrophages are characterized by low MHCII and high CD206.In contrast, anti-tumor M1 Macrophages are characterized by high MHCIIand low CD206. CT26 tumors from Formula C or vehicle-treated Balb/C micewith CT26 mouse colorectal cancer were analyzed for MHCII (FIG. 6B) andCD206 (FIG. 6C) levels 20 days after treatment started. The datademonstrated that Formula C-treated mice had higher MHCII levels andlower CD206 levels compared to vehicle-treated mice, indicating thatFormula C treatment shifts macrophages in vivo from pro-tumor (M2)macrophages to antitumor (M1) macrophages.

Example 4 Formula C Treatment Shifted CD8⁺ T Cells from Effector T Cellsto Memory T Cells

Spleens from the 2 BALB/C female mice (8.5 weeks old) were injected withcold PBS in a 6 mm plate to obtain cell suspension, using a 27 g needle.The spleens were then crushed using the back of a 10 ml syringe. Thesuspension was passed through a 21 g needle and filtered using a 70 umcells strainer. The plate was washed once with 10 ml PBS and thesolution was filtered as well. Cells were centrifuged for 10 min at 1200rpm, 4° C. The pellet was broken on the grill and re-suspended in 10 mlAKC X1 lysis buffer (1:10 dilution of X10 stock in Ultra-Pure Water).After 5 min on ice, PBS was added to a final volume of 50 ml, and cellswere centrifuged for 10 min at 1200 rpm, 4° C. Pellet was re-suspendedin 10 ml PBS and washed twice. After washing, pellet was re-suspended in20 ml DCs medium and passed through a 40 μm cell strainer. Cells weresuspended to 3×10⁶ ml with DC medium and supplemented with 1:1000 (100Units/ml) hIL-2 and 1:1000 (1 ug/ml) anti-CD28.

The cells were seeded 1 ml/well in a 24-well plate precoated with 5μg/ml 250 μl/well of anti-CD3e and carefully washed twice with 0.5 mlPBS (−/−), cells were added on the side of the wells to avoid washingaway the antibody.

Splenocytes were plated at 10⁶ cells/well in a 24-well plate coated with5 μg/ml anti-CD3e with soluble anti CD28 2 μg/ml and IL-2 100 units/ml.Cells were stimulated for 48 hours then reseeded 700,000 cells/well withthe different compounds for 24 hours in the presence of IL-1 100units/ml.

On the following day, cells were stained with: ghost dye 1 ul/sample(live/dead), and the following antibodies: CD62L, CD44, CD3, CD4, CD8and analyzed by fluorescence-activated cell sorting (FACS).

Naïve, non-stimulated cells demonstrated high CD62L and low CD44 (FIG.7A), while stimulated cells (vehicle, FIG. 7B, FIG. 7D) shifted to CD8+effector phenotype (lower right quadrant) and memory cell phenotype (topright quadrant). Formula C treatment shifted stimulated T cells tohigher CD44 and higher CD62L expression compared to vehicle-treated,signifying a shift from effector to memory cells (FIG. 7E). 2-DGtreatment had a similar effect to Formula C (FIG. 7C). Neither Formula Cnor 2-DG treatment significantly affected the percentage of live cells(FIG. 7F).

While certain features of the invention have been illustrated anddescribed herein, many modifications, substitutions, changes, andequivalents will now occur to those of ordinary skill in the art. It is,therefore, to be understood that the appended claims are intended tocover all such modifications and changes as fall within the true spiritof the invention.

What is claimed:
 1. A method of inducing an immune response orpotentiating an immune response in a subject, the method comprising thestep of contacting one or more cells of said subject with atherapeutically effective amount of a compound that promotes thedetachment of Hexokinase 2 (HK2) from the mitochondria, wherein saidcompound is represented by the structure of Formula (II):

wherein U and Z are each independently N or CH; X is O, NH, S, or abond; Y is CH₂, C═O, or C═S; R¹ is aryl, heteroaryl, or C(═O)—OR^(a),wherein aryl and heteroaryl are each optionally substituted with one ormore alkyl, arylalkyl, halogen, NO₂, CN, OR⁴, NR^(5a)R^(5b), or acombination thereof; R² and R³ are each independently at each occurrenceselected from the group consisting of: halogen, NO₂, CN, C₁-C₄ alkyl,OR⁴, and NR^(5a)R^(5b); R⁴, R^(a), R^(5a), and R^(5b) are eachindependently hydrogen or C₁-C₄ alkyl; n is 0 or 1; m is 0, 1, or 2; andp and q are each independently selected from 0, 1, 2, 3, and 4; or apharmaceutically acceptable salt thereof.
 2. The method of claim 1,wherein said immune response comprises conversion of macrophages from M2to M1 phenotype, activation of an NLRP inflammasome, or a combinationthereof.
 3. The method of claim 1, wherein the induction of said immuneresponse increases processing and/or secretion of interleukin (IL)-1β,IL-18, or a combination thereof.
 4. The method of claim 1, wherein saidimmune response comprises a cellular immune response.
 5. The method ofclaim 4, wherein said cellular immune response comprises activation ofT-cells, shifting effector T cells to memory T cells, activation of mastcells, lowering myeloid-derived suppressor cell (MDSC) levels,activation of dendritic cells or a combination thereof.
 6. The method ofclaim 5, wherein said dendritic cells are Langerin+dendritic cells. 7.The method of claim 1, wherein said subject suffers from ahyperproliferative disorder.
 8. The method of claim 7, wherein saidhyperproliferative disorder is a cancer, a tumor, a pre-cancerouscondition, or a benign hyperproliferative disorder.
 9. The method ofclaim 7, wherein said benign hyperproliferative disorder is LedderhoseDisease, Dupuytren's Disease, a keloid, or a hypertrophic scar.
 10. Themethod of claim 7, wherein said pre-cancerous condition is actinickeratosis or Bowen's Disease.
 11. The method of claim 7, wherein saidcancer is leukemia or a lymphoma.
 12. The method of claim 11, whereinsaid leukemia is acute myeloid leukemia (AML), acute lymphocyticleukemia (ALL), or B-cell chronic lymphocytic leukemia (B-CLL).
 13. Themethod of claim 11, wherein said lymphoma is a cutaneous T-cell lymphomaor a Non-Hodgkin lymphoma.
 14. The method of claim 13, wherein thecutaneous T-cell lymphoma is mycosis fungoides or Sezary syndrome. 15.The method of claim 7, wherein said cancer is a colon cancer, a stomachcancer, a head and neck cancer, a breast cancer, a prostate cancer, akidney cancer, a thyroid cancer, a skin cancer, a lung cancer, or apancreatic cancer.
 16. The method of claim 15, wherein said prostatecancer comprises metastatic castration resistant prostate cancer (CRPC).17. The method of claim 15, wherein said breast cancer comprises triplenegative breast cancer.
 18. The method of claim 15, wherein said lungcancer is a non-small cell lung cancer (NSCLC) or a small cell lungcancer (SCLC).
 19. The method of claim 15, wherein said skin cancer is amelanoma, a basal cell carcinoma or a squamous cell carcinoma.
 20. Themethod of claim 1, wherein said subject is immunocompromised.
 21. Themethod of claim 20, wherein said subject has an immunodeficiencydisorder.
 22. The method of claim 1, wherein said compound is present ina composition comprising a pharmaceutically acceptable carrier orexcipient.
 23. The method of claim 22, wherein said composition isformulated for parenteral administration or for oral, intravenous,subcutaneous, or intramuscular administration.
 24. The method of claim22, wherein said composition is formulated for topical administration.25. The method of claim 24, wherein said composition is formulated as anointment, a cream, a lotion, a foam, a paste, or a gel.
 26. The methodof claim 25, wherein said composition is formulated as a 5%, 10%, 20%,30%, 40% or 50% cream, gel, ointment or paste.
 27. The method accordingto claim 1, wherein said compound comprises:


28. The method according to claim 27, wherein said compound isformulated for oral administration.